JP2022028794A - Nanovesicles derived from morganella bacteria, and uses thereof - Google Patents

Nanovesicles derived from morganella bacteria, and uses thereof Download PDF

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JP2022028794A
JP2022028794A JP2021185327A JP2021185327A JP2022028794A JP 2022028794 A JP2022028794 A JP 2022028794A JP 2021185327 A JP2021185327 A JP 2021185327A JP 2021185327 A JP2021185327 A JP 2021185327A JP 2022028794 A JP2022028794 A JP 2022028794A
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キム、ユン－クン
Yoon-Keun Kim
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Abstract

PROBLEM TO BE SOLVED: To provide a method for diagnosing malignant diseases, such as gastric cancer, colon cancer, pancreatic cancer, bile duct cancer, breast cancer, ovarian cancer, bladder cancer, prostate cancer, and lymphoma; cardiovascular diseases, such as myocardial infarction, cardiomyopathy, atrial fibrillation, atypical angina, and stroke; diabetes; and Parkinson's disease, and also to provide a preventive or therapeutic composition for the above diseases or inflammatory diseases.
SOLUTION: A pharmaceutical composition comprising vesicles derived from bacteria of the genus Morganella as effective ingredients is used.
SELECTED DRAWING: Figure 23
COPYRIGHT: (C)2022,JPO&INPIT

Description

本発明は、モルガネラ属細菌由来ナノ小胞およびその用途に関し、より具体的には、モルガネラ属細菌に由来するナノ小胞を用いた胃癌、大腸癌、すい臓癌、胆管癌、乳癌、卵巣癌、膀胱癌、前立腺癌およびリンパ腫などの悪性疾患、心筋梗塞、心筋症、心房細動、異型狭心症、脳卒中などの心血管疾患、糖尿病、およびパーキンソン病の診断方法、および前記小胞を含む前記疾患または炎症性疾患に対する予防または治療用組成物に関する。 The present invention relates to nanovesicles derived from Morganella spp. And uses thereof, more specifically, gastric cancer, colon cancer, pancreatic cancer, bile duct cancer, breast cancer, ovarian cancer, using nanovesicles derived from Morganella spp. Methods for diagnosing malignant diseases such as bladder cancer, prostate cancer and lymphoma, cardiovascular diseases such as myocardial infarction, myocardial disease, atrial fibrillation, atypical angina, stroke, diabetes and Parkinson's disease, and said vesicles. Concerning preventive or therapeutic compositions for diseases or inflammatory diseases.

２１世紀に入って過去伝染病と認識された急性感染性疾患の重要性が低下することになる反面、ヒトとマイクロバイオームとの不調和によって発生する免疫機能の異常を伴った慢性疾患が生活の質と人間の寿命を決める主な疾患になり、疾病のパターンが変わった。２１世紀難治性慢性疾患として、癌、心血管疾患、慢性肺疾患、代謝疾患、炎症性疾患、および神経－精神疾患が、人間の寿命と生活の質を決める主な疾患として国民保健に大きい問題になっている。 While the importance of acute infectious diseases recognized as infectious diseases in the past will decrease in the 21st century, chronic diseases with abnormal immune function caused by incongruity between humans and microbiomes will become a part of daily life. It has become the main disease that determines quality and human lifespan, and the pattern of disease has changed. As intractable chronic diseases of the 21st century, cancer, cardiovascular disease, chronic lung disease, metabolic disease, inflammatory disease, and neuro-psychiatric disease are major problems for national health as the main diseases that determine human lifespan and quality of life. It has become.

炎症（Ｉｎｆｌａｍｍａｔｉｏｎ）は、細胞および組織の損傷や感染に対する局所的または全身的な防御機序であって、主に免疫系を成す体液性メディエーター（ｈｕｍｏｒａｌ ｍｅｄｉａｔｏｒ）が直接反応し、局所的または全身的作動システム（ｅｆｆｅｃｔｏｒ ｓｙｓｔｅｍ）を刺激することによって起こる連鎖的な生体反応により誘発される。主な炎症性疾患としては、胃炎、炎症性腸炎などの消化器疾患、歯周炎などの口腔疾患、喘息、慢性閉鎖性肺疾患（ＣＯＰＤ）、鼻炎などの呼吸器疾患、アトピー皮膚炎、脱毛、乾癬などの皮膚疾患、退行性関節炎、関節リウマチなどのような関節炎；および肥満、糖尿病、肝硬変症などの代謝疾患に含まれる。また、多様な研究を通じて持続的な炎症が癌を誘発することができるという結果が報告されてきた。 Inflammation is a local or systemic defense mechanism against cell and tissue damage and infection, with direct reaction by the humoral mediator, which primarily forms the immune system, and is local or systemic. It is triggered by a chain of biological reactions that occur by stimulating the effector system. The main inflammatory diseases are gastrointestinal diseases such as gastric inflammation and inflammatory arthritis, oral diseases such as periodontitis, asthma, chronic closed lung disease (COPD), respiratory diseases such as rhinitis, atopy dermatitis, and hair loss. , Skin disorders such as psoriasis, arthritis such as degenerative arthritis, rheumatoid arthritis; and metabolic disorders such as obesity, diabetes, liver cirrhosis. In addition, various studies have reported that persistent inflammation can induce cancer.

人体に共生する微生物は、１００兆に達し、ヒト細胞より１０倍多く、微生物の遺伝子数は、ヒト遺伝子数の１００倍を超えると知られている。微生物叢（ｍｉｃｒｏｂｉｏｔａあるいはｍｉｃｒｏｂｉｏｍｅ）は、与えられた棲息地に存在する真正細菌（ｂａｃｔｅｒｉａ）、古細菌（ａｒｃｈａｅａ）、真核生物（ｅｕｋａｒｙａ）を含む微生物群集（ｍｉｃｒｏｂｉａｌ ｃｏｍｍｕｎｉｔｙ）を言う。 It is known that the number of microorganisms that coexist in the human body reaches 100 trillion, which is 10 times more than that of human cells, and the number of genes of microorganisms exceeds 100 times that of human genes. The microbial flora (microbiota or microbiome) refers to a microbial community containing eubacteria, archaea, and eukarya that are present in a given habitat.

人体に共生する細菌および周辺環境に存在する細菌は、他の細胞への遺伝子、低分子化合物、タンパク質などの情報を交換するために、ナノメートルサイズの小胞（ｖｅｓｉｃｌｅ）を分泌する。粘膜は、２００ナノメートル（ｎｍ）サイズ以上の粒子は通過できない物理的な防御膜を形成して、粘膜に共生する細菌である場合には、粘膜を通過しないが、細菌由来小胞は、サイズが略１００ナノメートルサイズ以下であるので、比較的自由に粘膜を通じて上皮細胞を通過した後に人体に吸収される。局所的に分泌された細菌由来小胞は、粘膜の上皮細胞あるいは皮膚角質細胞を通じて吸収されて、局所炎症反応を誘導すると共に、人体に吸収されて各臓器に分布し、吸収された臓器で免疫および炎症反応を調節する。例えば、大腸菌（Ｅｓｈｃｈｅｒｉｃｈｉａ ｃｏｌｉ）のような病原性グラム陰性細菌に由来する小胞は、血管に吸収された場合に、血管内皮細胞の炎症反応を通じて全身的な炎症反応および血液凝固を促進させ、また、インスリンが作用する筋肉細胞などに吸収されて、インスリン抵抗性と糖尿病を誘発する。反面、有益な細菌に由来する小胞は、病原性小胞による免疫機能および代謝機能の異常を調節して疾病を調節することができる（Ｃｈｏｉ ＹＷ ｅｔ ａｌ．，Ｇｕｔ ｍｉｃｒｏｂｅ－ｄｅｒｉｖｅｄ ｅｘｔｒａｃｅｌｌｕｌａｒ ｖｅｓｉｃｌｅｓ ｉｎｄｕｃｅ ｉｎｓｕｌｉｎ ｒｅｓｉｓｔａｎｃｅ，ｔｈｅｒｅｂｙ ｉｍｐａｉｒｉｎｇ ｇｌｕｃｏｓｅ ｍｅｔａｂｏｌｉｓｍ ｉｎ ｓｋｅｌｅｔａｌ ｍｕｓｃｌｅ．Ｓｃｉｅｎｔｉｆｉｃ Ｒｅｐｏｒｔｓ，２０１５．）。 Bacteria that coexist in the human body and those that exist in the surrounding environment secrete nanometer-sized vesicles to exchange information such as genes, small molecule compounds, and proteins with other cells. The mucosa forms a physical protective membrane that particles larger than 200 nanometers (nm) in size cannot pass through, and if it is a bacterium that coexists with the mucosa, it does not pass through the mucosa, but bacterial-derived vesicles are sized. Is less than about 100 nanometers in size, so it is relatively freely absorbed by the human body after passing through epithelial cells through the mucosa. Locally secreted bacterial-derived vesicles are absorbed through mucosal epithelial cells or cutaneous keratinocytes to induce a local inflammatory reaction, and are absorbed by the human body and distributed to each organ, and are immunized by the absorbed organs. And regulate the inflammatory response. For example, vesicles derived from pathogenic gram-negative bacteria such as Eshchericia coli promote systemic inflammatory response and blood coagulation through the inflammatory response of vascular endothelial cells when absorbed into blood vessels. , It is absorbed by muscle cells on which insulin acts and induces insulin resistance and diabetes. On the other hand, vesicles derived from beneficial bacteria can regulate diseases by regulating abnormalities in immune and metabolic functions caused by pathogenic vesicles (Choi YW et al., Gut microbe-developed extracellular metabolism insulin). response, thereby impairing glucose metabolism in skeletal muscle. Scientific Reports, 2015.).

細菌に由来する小胞などの因子に対する免疫反応は、ＩＬ－１７サイトカインの分泌を特徴とするＴｈ１７免疫反応であって、これは、細菌由来小胞に曝露時にＩＬ－６が分泌され、これは、Ｔｈ１７免疫反応を誘導する。Ｔｈ１７免疫反応による炎症は、好中球の浸潤を特徴とし、炎症が発生する過程でマクロファージなどのような炎症細胞で分泌されるＴＮＦ－ａｌｐｈａが重要な役割を担当する。 The immune response to factors such as bacterial vesicles is a Th17 immune response characterized by the secretion of IL-17 cytokines, which secrete IL-6 upon exposure to bacterial vesicles. , Th17 Induces an immune response. Inflammation due to the Th17 immune response is characterized by infiltration of neutrophils, and TNF-alpha secreted by inflammatory cells such as macrophages plays an important role in the process of inflammation development.

モルガネラ属細菌は、嫌気性グラム陰性細菌であって、人間と動物の腸に共生する細菌と知られている。前記細菌のうちモルガネラ・モルガニイ（Ｍｏｒｇａｎｅｌｌａ ｍｏｒｇａｎｉｉ）菌は、手術後に感染または尿道感染などを起こす病原性細菌と知られている。しかしながら、現在まで、モルガネラ属細菌が細胞外に小胞を分泌するという事実は報告されておらず、特に癌、心血管疾患、代謝疾患、炎症性疾患、および神経－精神疾患の診断および治療に応用した事例は報告されたことがない。 Morganella bacteria are anaerobic gram-negative bacteria and are known to coexist in the intestines of humans and animals. Among the above-mentioned bacteria, Morganella morganii is known as a pathogenic bacterium that causes infection or urinary tract infection after surgery. However, to date, the fact that Morganella bacteria secrete extracellular vesicles has not been reported, especially for the diagnosis and treatment of cancer, cardiovascular disease, metabolic disease, inflammatory disease, and neuro-psychiatric disorders. No application cases have been reported.

これより、本発明では、モルガネラ属細菌由来小胞が正常ヒトに比べて癌、心血管疾患、代謝疾患、炎症性疾患、および神経－精神疾患の患者の臨床サンプルに有意に減少していることを確認して、疾病を診断することができることを確認した。また、モルガネラ・モルガニイ菌から小胞を分離し、特性を分析した結果、悪性疾患、心血管疾患、代謝疾患、炎症性疾患、および神経－精神疾患の予防または治療用組成物に用いられることを確認した。 Thus, in the present invention, Morganella-derived vesicles are significantly reduced in clinical samples of patients with cancer, cardiovascular disease, metabolic disease, inflammatory disease, and neuro-psychiatric disease compared to normal humans. Confirmed that the disease can be diagnosed. In addition, as a result of isolating vesicles from Morganella and Morganii bacteria and analyzing their characteristics, they have been found to be used in compositions for the prevention or treatment of malignant diseases, cardiovascular diseases, metabolic diseases, inflammatory diseases, and neuro-psychiatric diseases. confirmed.

本発明者らは、前記のような従来の問題点を解決するために鋭意研究した結果、メタゲノム分析を通して正常ヒトに比べて胃癌、大腸癌、すい臓癌、胆管癌、乳癌、卵巣癌、膀胱癌、前立腺癌およびリンパ腫などの悪性疾患、心筋梗塞、心筋症、心房細動、異型狭心症、脳卒中などの心血管疾患、糖尿病、およびパーキンソン病などの患者臨床サンプルでモルガネラ属細菌由来小胞の含量が有意に減少していることを確認した。また、モルガネラ属細菌に属するモルガネラ・モルガニイ菌から小胞を分離してマクロファージに処理したとき、病原性小胞によるＴＮＦ－ａｌｐｈａの分泌を顕著に抑制することを確認し、これに基づいて本発明を完成した。 As a result of diligent research to solve the above-mentioned conventional problems, the present inventors have obtained gastric cancer, colon cancer, pancreatic cancer, bile duct cancer, breast cancer, ovarian cancer, and bladder cancer as compared with normal humans through metagenome analysis. Malignant diseases such as prostate cancer and lymphoma, myocardial infarction, myocardial disease, atrial fibrillation, atypical angina, cardiovascular diseases such as stroke, diabetes, and patients with clinical samples such as Parkinson's disease. It was confirmed that the content was significantly reduced. Further, it was confirmed that when vesicles were isolated from Morganella-Morganella bacterium belonging to the genus Morganella and treated into macrophages, the secretion of TNF-alpha by pathogenic vesicles was remarkably suppressed, and the present invention was based on this. Was completed.

これより、本発明は、胃癌、大腸癌、すい臓癌、胆管癌、乳癌、卵巣癌、膀胱癌、前立腺癌、リンパ腫、心筋梗塞、心筋症、心房細動、異型狭心症、脳卒中、糖尿病、またはパーキンソン病の診断のための情報提供方法を提供することを目的とする。 Thus, the present invention relates to gastric cancer, colon cancer, pancreatic cancer, bile duct cancer, breast cancer, ovarian cancer, bladder cancer, prostate cancer, lymphoma, myocardial infarction, myocardial infarction, atrial fibrillation, atypical angina, stroke, diabetes, Or, the purpose is to provide a method of providing information for diagnosing Parkinson's disease.

また、本発明は、モルガネラ由来小胞を有効成分として含む胃癌、大腸癌、すい臓癌、胆管癌、乳癌、卵巣癌、膀胱癌、前立腺癌、リンパ腫、心筋梗塞、心筋症、心房細動、異型狭心症、脳卒中、糖尿病、パーキンソン病、または炎症性疾患の予防または治療用組成物を提供することを他の目的とする。 In addition, the present invention also comprises gastric cancer, colon cancer, pancreatic cancer, bile duct cancer, breast cancer, ovarian cancer, bladder cancer, prostate cancer, lymphoma, myocardial infarction, myocardial disease, atrial fibrillation, and atypia containing Morganella-derived vesicles as an active ingredient. Other objectives are to provide a composition for the prevention or treatment of angina, stroke, diabetes, Parkinson's disease, or inflammatory disease.

しかしながら、本発明が解決しようとする技術的課題は、以上で言及した課題に制限されず、言及されていない他の課題は、下記の記載から当業者に明確に理解され得る。 However, the technical problem to be solved by the present invention is not limited to the problem mentioned above, and other problems not mentioned above can be clearly understood by those skilled in the art from the following description.

前記のような本発明の目的を達成するために、本発明は、下記の段階を含む、胃癌、大腸癌、すい臓癌、胆管癌、乳癌、卵巣癌、膀胱癌、前立腺癌、リンパ腫、心筋梗塞、心筋症、心房細動、異型狭心症、脳卒中、糖尿病、またはパーキンソン病の診断のための情報提供方法を提供する：
（ａ）正常ヒトおよび被検者のサンプルから分離した小胞からＤＮＡを抽出する段階；
（ｂ）前記抽出したＤＮＡに対して１６Ｓ ｒＤＮＡに存在する遺伝子配列に基づいて作製したプライマーペアを用いてＰＣＲを行って、それぞれのＰＣＲ産物を収得する段階；および
（ｃ）前記ＰＣＲ産物の定量分析を通して正常ヒトに比べてモルガネラ属細菌由来小胞の含量が低い場合、胃癌、大腸癌、すい臓癌、胆管癌、乳癌、卵巣癌、膀胱癌、前立腺癌、リンパ腫、心筋梗塞、心筋症、心房細動、異型狭心症、脳卒中、糖尿病、またはパーキンソン病と判定する段階。 In order to achieve the above-mentioned object of the present invention, the present invention includes gastric cancer, colon cancer, pancreatic cancer, bile duct cancer, breast cancer, ovarian cancer, bladder cancer, prostate cancer, lymphoma, myocardial infarction, including the following stages. , Provides informational methods for the diagnosis of myocardial infarction, atrial fibrillation, atypical angina, stroke, diabetes, or Parkinson's disease:
(A) Step of extracting DNA from vesicles isolated from normal human and subject samples;
(B) Steps of obtaining each PCR product by performing PCR on the extracted DNA using a primer pair prepared based on the gene sequence present in 16S rDNA; and (c) Quantification of the PCR product. When the content of vesicles derived from Morganella spp. The stage of determining fibrillation, atypical angina, stroke, diabetes, or Parkinson's disease.

また、本発明は、下記の段階を含む、胃癌、大腸癌、すい臓癌、胆管癌、乳癌、卵巣癌、膀胱癌、前立腺癌、リンパ腫、心筋梗塞、心筋症、心房細動、異型狭心症、脳卒中、糖尿病、またはパーキンソン病の診断方法を提供する：
（ａ）正常ヒトおよび被検者のサンプルから分離した小胞からＤＮＡを抽出する段階；
（ｂ）前記抽出したＤＮＡに対して１６Ｓ ｒＤＮＡに存在する遺伝子配列に基づいて作製したプライマーペアを用いてＰＣＲを行って、それぞれのＰＣＲ産物を収得する段階；および
（ｃ）前記ＰＣＲ産物の定量分析を通して正常ヒトに比べてモルガネラ属細菌由来小胞の含量が低い場合、胃癌、大腸癌、すい臓癌、胆管癌、乳癌、卵巣癌、膀胱癌、前立腺癌、リンパ腫、心筋梗塞、心筋症、心房細動、異型狭心症、脳卒中、糖尿病、またはパーキンソン病と判定する段階。 The present invention also includes gastric cancer, colon cancer, pancreatic cancer, bile duct cancer, breast cancer, ovarian cancer, bladder cancer, prostate cancer, lymphoma, myocardial infarction, myocardial disease, atrial fibrillation, and atypical angina. , Provides methods for diagnosing stroke, diabetes, or Parkinson's disease:
(A) Step of extracting DNA from vesicles isolated from normal human and subject samples;
(B) Steps of obtaining each PCR product by performing PCR on the extracted DNA using a primer pair prepared based on the gene sequence present in 16S rDNA; and (c) Quantification of the PCR product. When the content of vesicles derived from Morganella spp. The stage of determining fibrillation, atypical angina, stroke, diabetes, or Parkinson's disease.

本発明のさらに他の具現例において、前記段階（ａ）でのサンプルは、血液、尿、または便でありうる。 In yet another embodiment of the invention, the sample in step (a) can be blood, urine, or stool.

本発明のさらに他の具現例において、前記段階（ｂ）でのプライマーペアは、配列番号１および配列番号２のプライマーでありうる。 In still another embodiment of the invention, the primer pair in step (b) can be the primers of SEQ ID NO: 1 and SEQ ID NO: 2.

また、本発明は、モルガネラ属細菌由来小胞を有効成分として含む、胃癌、大腸癌、すい臓癌、胆管癌、乳癌、卵巣癌、膀胱癌、前立腺癌、リンパ腫、心筋梗塞、心筋症、心房細動、異型狭心症、脳卒中、糖尿病、パーキンソン病、および炎症性疾患よりなる群から選ばれる１つ以上の疾病の予防または治療用薬学的組成物を提供する。 In addition, the present invention also contains gastric cancer, colon cancer, pancreatic cancer, bile duct cancer, breast cancer, ovarian cancer, bladder cancer, prostate cancer, lymphoma, myocardial infarction, myocardial disease, and atrial appendage, which contain vesicles derived from Morganella spp. Provided are pharmaceutical compositions for the prevention or treatment of one or more diseases selected from the group consisting of movement, atypical angina, stroke, diabetes, Parkinson's disease, and inflammatory diseases.

また、本発明は、モルガネラ属細菌由来小胞を有効成分として含む、胃癌、大腸癌、すい臓癌、胆管癌、乳癌、卵巣癌、膀胱癌、前立腺癌、リンパ腫、心筋梗塞、心筋症、心房細動、異型狭心症、脳卒中、糖尿病、パーキンソン病、および炎症性疾患よりなる群から選ばれる１つ以上の疾病の予防または改善用食品組成物を提供する。 In addition, the present invention also contains gastric cancer, colon cancer, pancreatic cancer, bile duct cancer, breast cancer, ovarian cancer, bladder cancer, prostate cancer, lymphoma, myocardial infarction, myocardial disease, and atrial appendage, which contain vesicles derived from Morganella spp. Provided are food compositions for the prevention or amelioration of one or more diseases selected from the group consisting of movement, atypical angina, stroke, diabetes, Parkinson's disease, and inflammatory diseases.

また、本発明は、モルガネラ属細菌由来小胞を有効成分として含む薬学的組成物を個体に投与する段階を含む、胃癌、大腸癌、すい臓癌、胆管癌、乳癌、卵巣癌、膀胱癌、前立腺癌、リンパ腫、心筋梗塞、心筋症、心房細動、異型狭心症、脳卒中、糖尿病、パーキンソン病、および炎症性疾患よりなる群から選ばれる１つ以上の疾病の予防または治療方法を提供する。 The present invention also includes gastric cancer, colon cancer, pancreatic cancer, bile duct cancer, breast cancer, ovarian cancer, bladder cancer, and prostate, which comprises the step of administering to an individual a pharmaceutical composition containing vesicles derived from Morganella spp. Provided are methods for preventing or treating one or more diseases selected from the group consisting of cancer, lymphoma, myocardial infarction, myocardial disease, atrial fibrillation, atypical angina, stroke, diabetes, Parkinson's disease, and inflammatory disease.

また、本発明は、モルガネラ属細菌由来小胞の、胃癌、大腸癌、すい臓癌、胆管癌、乳癌、卵巣癌、膀胱癌、前立腺癌、リンパ腫、心筋梗塞、心筋症、心房細動、異型狭心症、脳卒中、糖尿病、パーキンソン病、および炎症性疾患よりなる群から選ばれる１つ以上の疾病の予防または治療用途を提供する。 The present invention also relates to gastric cancer, colon cancer, pancreatic cancer, bile duct cancer, breast cancer, ovarian cancer, bladder cancer, prostate cancer, lymphoma, myocardial infarction, myocardial disease, atrial fibrillation, and atypical narrowing of vesicles derived from Morganella spp. It provides a prophylactic or therapeutic use for one or more diseases selected from the group consisting of heart disease, stroke, diabetes, Parkinson's disease, and inflammatory diseases.

本発明の一具現例において、前記炎症性疾患は、アトピー皮膚炎、にきび、乾癬、副鼻腔炎、鼻炎、結膜炎、喘息、皮膚炎、炎症性コラーゲン血管疾患、糸球体腎炎、脳炎、炎症性腸炎、慢性閉鎖性肺疾患、敗血症、敗血症性ショック、肺線維症、未分化脊椎関節症、未分化関節症、関節炎、炎症性骨溶解、ウイルスまたはバクテリア感染による慢性炎症疾患、大腸炎、潰瘍性大腸炎、炎症性腸疾患、関節炎、関節リウマチ、反応性関節炎、骨関節炎、鞏皮症、骨粗しょう症、アテローム性動脈硬化症、心筋炎、心内膜炎、心嚢炎、嚢胞性線維症、橋本甲状腺炎、グレーブス病、ハンセン病、梅毒、ライム病（Ｌｙｍｅ ｄｉｓｅａｓｅ）、ボレリア症（Ｂｏｒｒｅｌｉｏｓｉｓ）、神経性ボレリア症、結核、サルコイドーシス（Ｓａｒｃｏｉｄｏｓｉｓ）、ループス、凍瘡状ループス、結核性ループス、ループス腎炎、全身性エリテマトーデス、黄斑変性、ブドウ膜炎、過敏性腸症候群、クローン病、シェーグレン症候群、線維筋痛、慢性疲労症候群、慢性疲労免疫不全症候群、筋痛性脳脊髄炎、筋萎縮性側索硬化症、パーキンソン病、および多発性硬化症よりなる群から選ばれる１つ以上でありうる。 In one embodiment of the present invention, the inflammatory disease is atopy dermatitis, acne, psoriasis, sinusitis, rhinitis, conjunctivitis, asthma, dermatitis, inflammatory collagen vascular disease, glomerular nephritis, encephalitis, inflammatory enteritis. , Chronic closed lung disease, septicemia, septic shock, pulmonary fibrosis, undifferentiated lupus erythematosus, undifferentiated arthritis, arthritis, inflammatory osteolysis, chronic inflammatory disease due to viral or bacterial infection, colitis, ulcerative colon Flame, inflammatory bowel disease, arthritis, rheumatoid arthritis, reactive arthritis, osteoarthritis, lupus erythematosus, osteoporosis, atherosclerosis, myocarditis, endocarditis, cystitis, cystic fibrosis, Hashimoto Lupus erythematosus, Graves' disease, Hansen's disease, syphilis, Lyme disease, Borreliosis, neuroborreliosis, tuberculosis, sarcoidosis, lupus, lupus erythematosus, tuberculous lupus, lupus nephritis, systemic Lupus erythematosus, lupus erythematosus, vegetationitis, irritable bowel syndrome, Crohn's disease, Schegren's syndrome, fibromyalgia, chronic fatigue syndrome, chronic fatigue immunodeficiency syndrome, myopathic encephalomyelitis, muscular atrophic lateral sclerosis, Parkinson It can be one or more selected from the group consisting of disease and multiple sclerosis.

本発明の他の具現例において、前記炎症性疾患は、ＩＬ－６またはＴＮＦ－αにより媒介される疾患でありうる。 In another embodiment of the invention, the inflammatory disease can be a disease mediated by IL-6 or TNF-α.

本発明のさらに他の具現例において、前記小胞は、平均直径が１０～２００ｎｍのものでありうる。 In yet another embodiment of the invention, the vesicles may have an average diameter of 10-200 nm.

本発明のさらに他の具現例において、前記小胞は、モルガネラ属細菌から自然的にまたは人工的に分泌されるものでありうる。 In yet another embodiment of the invention, the vesicles may be naturally or artificially secreted by a Morganella bacterium.

本発明のさらに他の具現例において、前記モルガネラ属細菌由来小胞は、モルガネラ・モルガニイから分泌されるものでありうる。 In still another embodiment of the invention, the Morganella bacterium-derived vesicles may be secreted from Morganella morganii.

本発明者らは、腸内細菌である場合には、体内に吸収されないが、細菌由来小胞である場合には、上皮細胞を通じて体内に吸収されて、全身的に分布し、腎臓、肝臓、肺を通じて体外に排泄されることを確認し、患者の血液、尿、または便などに存在する細菌由来小胞メタゲノム分析を通して胃癌、大腸癌、すい臓癌、胆管癌、乳癌、卵巣癌、膀胱癌、前立腺癌、リンパ腫、心筋梗塞、心筋症、心房細動、異型狭心症、脳卒中、糖尿病、およびパーキンソン病の患者の血液、尿、または便に存在するモルガネラ属細菌由来小胞が正常ヒトに比べて有意に減少していることを確認した。また、モルガネラ属細菌の一種であるモルガネラ・モルガニイを体外で培養して小胞を分離して、体外で炎症細胞に投与したとき、病原性小胞による炎症メディエーターの分泌を有意に抑制すると同時に、癌動物モデルで癌の発生を抑制することを観察したところ、本発明によるモルガネラ属細菌由来小胞は、胃癌、大腸癌、すい臓癌、胆管癌、乳癌、卵巣癌、膀胱癌、前立腺癌、リンパ腫、心筋梗塞、心筋症、心房細動、異型狭心症、脳卒中、糖尿病、およびパーキンソン病に対する診断方法、および前記疾患または炎症性疾患に対する食品または薬物などの予防用あるいは治療用組成物に有用に用いられるものと期待される。 In the case of enterobacteria, the present inventors are not absorbed into the body, but in the case of bacterial-derived vesicles, they are absorbed into the body through epithelial cells and distributed systemically, and the kidneys, liver, etc. Confirmed to be excreted from the body through the lungs, and through analysis of bacterial vesicle metagenome present in the patient's blood, urine, or stool, gastric cancer, colon cancer, pancreatic cancer, bile duct cancer, breast cancer, ovarian cancer, bladder cancer, Compared to normal humans, Morganella-derived follicles present in the blood, urine, or stool of patients with prostate cancer, lymphoma, myocardial infarction, myocardial disease, atrial fibrillation, atypical angina, stroke, diabetes, and Parkinson's disease It was confirmed that there was a significant decrease. In addition, when Morganella morganii, a type of Morganella genus bacterium, is cultured in vitro to isolate vesicles and administered to inflammatory cells in vitro, the secretion of inflammatory mediators by pathogenic vesicles is significantly suppressed, and at the same time. When it was observed to suppress the development of cancer in a cancer animal model, the vesicles derived from Morganella spp. According to the present invention were gastric cancer, colon cancer, pancreatic cancer, bile duct cancer, breast cancer, ovarian cancer, bladder cancer, prostate cancer, lymphoma. , A diagnostic method for myocardial infarction, myocardial disease, atrial fibrillation, atypical angina, stroke, diabetes, and Parkinson's disease, and prophylactic or therapeutic compositions such as foods or drugs for said or inflammatory diseases. Expected to be used.

図１ａは、マウスに細菌と細菌由来小胞（ＥＶ）を口腔で投与した後、時間別に細菌と小胞の分布様相を撮影した写真であり、図１ｂは、口腔で投与した後１２時間目に、血液、腎臓、肝臓、および様々な臓器を摘出して、細菌と小胞の体内分布様相を評価した結果である。FIG. 1a is a photograph of the distribution of bacteria and vesicles taken by time after oral administration of bacteria and vesicles derived from bacteria (EV) to mice, and FIG. 1b is a photograph taken 12 hours after oral administration. It is the result of extracting blood, kidney, liver, and various organs and evaluating the distribution of bacteria and vesicles in the body. 図２ａ～図２ｃは、胃癌患者および正常ヒトの便（図２ａ）、血液（図２ｂ）、および尿（図２ｃ）に存在する細菌由来小胞メタゲノム分析を実施した後、モルガネラ属細菌由来小胞の分布を比較した結果である。2a-2c show bacterial-derived vesicle metagenome analysis present in stool (FIG. 2a), blood (FIG. 2b), and urine (FIG. 2c) of gastric cancer patients and normal humans, followed by small Morganella-derived bacteria. It is the result of comparing the distribution of vesicles. 図３は、大腸癌患者および正常ヒトの尿に存在する細菌由来小胞メタゲノム分析を実施した後、モルガネラ属細菌由来小胞の分布を比較した結果である。FIG. 3 shows the results of comparing the distribution of vesicles derived from Morganella spp. After performing metagenome analysis of bacterial vesicles present in the urine of colorectal cancer patients and normal humans. 図４は、すい臓癌患者および正常ヒトの血液に存在する細菌由来小胞メタゲノム分析を実施した後、モルガネラ属細菌由来小胞の分布を比較した結果である。FIG. 4 shows the results of comparing the distribution of vesicles derived from Morganella spp. After performing metagenome analysis of bacterial vesicles present in the blood of patients with pancreatic cancer and normal humans. 図５は、胆管癌患者および正常ヒトの血液に存在する細菌由来小胞メタゲノム分析を実施した後、モルガネラ属細菌由来小胞の分布を比較した結果である。FIG. 5 shows the results of comparing the distribution of vesicles derived from Morganella spp. After performing metagenome analysis of bacterial vesicles present in the blood of patients with bile duct cancer and normal humans. 図６は、乳癌患者および正常ヒト尿に存在する細菌由来小胞メタゲノム分析を実施した後、モルガネラ属細菌由来小胞の分布を比較した結果である。FIG. 6 shows the results of comparing the distribution of vesicles derived from Morganella spp. After performing metagenome analysis of vesicles derived from bacteria present in breast cancer patients and normal human urine. 図７ａおよび図７ｂは、卵巣癌患者および正常ヒトの血液（図７ａ）および尿（図７ｂ）に存在する細菌由来小胞メタゲノム分析を実施した後、モルガネラ属細菌由来小胞の分布を比較した結果である。7a and 7b compare the distribution of bacterial vesicles of the genus Morganella after performing a metagenome analysis of bacterial vesicles present in blood (FIG. 7a) and urine (FIG. 7b) of ovarian cancer patients and normal humans. The result. 図８ａおよび図８ｂは、膀胱癌患者および正常ヒトの血液（図８ａ）および尿（図８ｂ）に存在する細菌由来小胞メタゲノム分析を実施した後、モルガネラ属細菌由来小胞の分布を比較した結果である。8a and 8b compare the distribution of bacterial vesicles derived from Morganella after performing a metagenome analysis of bacterial vesicles present in blood (FIG. 8a) and urine (FIG. 8b) of bladder cancer patients and normal humans. The result. 図９は、前立腺癌患者および正常ヒトの尿に存在する細菌由来小胞メタゲノム分析を実施した後、モルガネラ属細菌由来小胞の分布を比較した結果である。FIG. 9 shows the results of comparing the distribution of vesicles derived from Morganella spp. After performing metagenome analysis of bacterial vesicles present in the urine of prostate cancer patients and normal humans. 図１０は、リンパ腫患者および正常ヒトの血液に存在する細菌由来小胞メタゲノム分析を実施した後、モルガネラ属細菌由来小胞の分布を比較した結果である。FIG. 10 shows the results of comparing the distribution of vesicles derived from Morganella spp. After performing metagenome analysis of bacterial vesicles present in the blood of lymphoma patients and normal humans. 図１１は、心筋梗塞患者および正常ヒトの血液に存在する細菌由来小胞メタゲノム分析を実施した後、モルガネラ属細菌由来小胞の分布を比較した結果である。FIG. 11 shows the results of comparing the distribution of vesicles derived from Morganella spp. After performing metagenome analysis of bacterial vesicles present in the blood of patients with myocardial infarction and normal humans. 図１２は、心筋症患者および正常ヒトの血液に存在する細菌由来小胞メタゲノム分析を実施した後、モルガネラ属細菌由来小胞の分布を比較した結果である。FIG. 12 shows the results of comparing the distribution of vesicles derived from Morganella spp. After performing metagenome analysis of bacterial vesicles present in the blood of cardiomyopathy patients and normal humans. 図１３は、心房細動患者および正常ヒトの血液に存在する細菌由来小胞メタゲノム分析を実施した後、モルガネラ属細菌由来小胞の分布を比較した結果である。FIG. 13 shows the results of comparing the distribution of bacterial vesicles of the genus Morganella after performing a metagenome analysis of bacterial vesicles present in the blood of patients with atrial fibrillation and normal humans. 図１４は、異型狭心症患者および正常ヒトの血液に存在する細菌由来小胞メタゲノム分析を実施した後、モルガネラ属細菌由来小胞の分布を比較した結果である。FIG. 14 shows the results of comparing the distribution of bacterial vesicles of the genus Morganella after performing a metagenome analysis of bacterial vesicles present in the blood of patients with atypical angina and normal humans. 図１５は、脳卒中患者および正常ヒトの血液に存在する細菌由来小胞メタゲノム分析を実施した後、モルガネラ属細菌由来小胞の分布を比較した結果である。FIG. 15 shows the results of comparing the distribution of bacterial vesicles of the genus Morganella after performing a metagenome analysis of bacterial vesicles present in the blood of stroke patients and normal humans. 図１６ａおよび図１６ｂは、糖尿病患者および正常ヒトの血液（図１６ａ）および尿（図１６ｂ）に存在する細菌由来小胞メタゲノム分析を実施した後、モルガネラ属細菌由来小胞の分布を比較した結果である。16a and 16b show the results of comparing the distribution of bacterial vesicles of the genus Morganella after performing a metagenome analysis of bacterial vesicles present in blood (FIG. 16a) and urine (FIG. 16b) of diabetic patients and normal humans. Is. 図１７は、パーキンソン病患者および正常ヒトの尿に存在する細菌由来小胞メタゲノム分析を実施した後、モルガネラ属細菌由来小胞の分布を比較した結果である。FIG. 17 shows the results of comparing the distribution of vesicles derived from Morganella spp. After performing metagenome analysis of bacterial vesicles present in the urine of Parkinson's disease patients and normal humans. 図１８は、モルガネラ・モルガニイ由来小胞の細胞死滅効果を評価するために、モルガネラ・モルガニイ由来小胞をマクロファージに処理して、モルガネラ・モルガニイ由来小胞の細胞死滅効果を評価した結果である。FIG. 18 shows the results of treating Morganella-Morganella-derived vesicles with macrophages to evaluate the cell-killing effect of Morganella-Morganella-derived vesicles in order to evaluate the cell-killing effect of Morganella-Morganella-derived vesicles. 図１９ａおよび図１９ｂは、モルガネラ・モルガニイ由来小胞の抗炎症効果を評価するために、病原性小胞である大腸菌小胞（Ｅ．ｃｏｌｉ ＥＶ）の処理前にモルガネラ・モルガニイ由来小胞を前処理して、大腸菌小胞による炎症メディエーターであるＩＬ－６（図１９ａ）およびＴＮＦ－α（図１９ｂ）の分泌に及ぼす影響を評価した結果である。19a and 19b show Morganella-Morganii-derived vesicles prior to treatment with the pathogenic E. coli EV to evaluate the anti-inflammatory effect of Morganella-Morganii-derived vesicles. It is the result of processing and evaluating the influence on the secretion of the inflammation mediators IL-6 (FIG. 19a) and TNF-α (FIG. 19b) by E. coli vesicles. 図２０は、モルガネラ・モルガニイ由来小胞の抗炎症効果に対する互いに異なる菌株に由来した小胞の影響を比較するために、病原性小胞である大腸菌小胞（Ｅ．ｃｏｌｉ ＥＶ）の処理前に互いに異なるヒトから分離したモルガネラ・モルガニイ（ＭＭＲ１０１、ＭＭＲ２０１）由来小胞を前処理して、大腸菌小胞によるＴＮＦ－αの分泌に及ぼす影響を評価した結果である（ＮＣ：ｎｅｇａｔｉｖｅ ｃｏｎｔｒｏｌ；ＰＣ：ｐｏｓｉｔｉｖｅ ｃｏｎｔｒｏｌ；Ｌ．ｐｌａｎｔａｒｕｍ：Ｌａｃｔｏｂａｃｉｌｌｕｓ ｐｌａｎｔａｒｕｍ）。FIG. 20 shows the effect of vesicles derived from different strains on the anti-inflammatory effect of vesicles derived from Morganella morganii prior to treatment with pathogenic E. coli EV. This is the result of pretreating vesicles derived from Morganella morganii (MMR101, MMR201) isolated from different humans and evaluating the effect of Escherichia coli vesicles on the secretion of TNF-α (NC: negative control; PC: pathogenic). control; L. colitraum: Lactobacillus plantarum). 図２１は、モルガネラ・モルガニイ由来小胞の抗炎症効果に対する熱処理または酸処理の影響を評価するために、病原性小胞である大腸菌小胞（Ｅ．ｃｏｌｉ ＥＶ）の処理前に熱処理または酸処理したモルガネラ・モルガニイ（ＭＭＲ１０１，ＭＭＲ２０１）由来小胞を前処理して、大腸菌小胞によるＴＮＦ－αの分泌に及ぼす影響を評価した結果である（ＮＣ：ｎｅｇａｔｉｖｅ ｃｏｎｔｒｏｌ；ＰＣ：ｐｏｓｉｔｉｖｅ ｃｏｎｔｒｏｌ；Ｌ．ｐｌａｎｔａｒｕｍ：Ｌａｃｔｏｂａｃｉｌｌｕｓ ｐｌａｎｔａｒｕｍ）。FIG. 21 shows the effect of heat treatment or acid treatment on the anti-inflammatory effect of Morganella morganii-derived vesicles prior to treatment of pathogenic E. coli vesicles (E. coli EV). This is the result of pretreating vesicles derived from Morganella morganii (MMR101, MMR201) and evaluating the effect of Escherichia coli vesicles on the secretion of TNF-α (NC: negative control; PC: positive control; L. plantarum). : Lactobacillus plantarum). 図２２は、モルガネラ・モルガニイ由来小胞の抗癌効能を評価するために、モルガネラ・モルガニイ由来小胞をマウスに投与したプロトコルである。FIG. 22 is a protocol in which Morganella-Morganella-derived vesicles were administered to mice to evaluate the anti-cancer efficacy of Morganella-Morganella-derived vesicles. 図２３は、モルガネラ・モルガニイ由来小胞の抗癌効能を評価するために、モルガネラ・モルガニイ小胞を腹腔（ＩＰ）または経口（ＰＯ）で投与して、癌細胞による腫瘍発生に及ぼす影響を評価した結果である。In FIG. 23, in order to evaluate the anticancer efficacy of Morganella-Morganella-derived vesicles, Morganella-Morganella vesicles are administered intraperitoneally (IP) or orally (PO) to evaluate the effect on tumorigenesis by cancer cells. Is the result of this.

本発明は、モルガネラ属細菌由来小胞およびその用途に関する。 The present invention relates to vesicles derived from Morganella spp. And uses thereof.

本発明者らは、メタゲノム分析を通して正常ヒトに比べて胃癌、大腸癌、すい臓癌、胆管癌、乳癌、卵巣癌、膀胱癌、前立腺癌、リンパ腫、心筋梗塞、心筋症、心房細動、異型狭心症、脳卒中、糖尿病、およびパーキンソン病の患者由来サンプルでモルガネラ属細菌由来小胞の含量が顕著に減少していることを確認したところ、これに基づいて本発明を完成した。 Through metagenome analysis, the present inventors have gastric cancer, colon cancer, pancreatic cancer, bile duct cancer, breast cancer, ovarian cancer, bladder cancer, prostate cancer, lymphoma, myocardial infarction, myocardial disease, atrial fibrillation, and atypical narrowing. It was confirmed that the content of vesicles derived from Morganella spp. Was significantly reduced in the samples derived from patients with heart disease, stroke, diabetes, and Parkinson's disease, and the present invention was completed based on this.

これより、本発明は、下記の段階を含む、胃癌、大腸癌、すい臓癌、胆管癌、乳癌、卵巣癌、膀胱癌、前立腺癌、リンパ腫、心筋梗塞、心筋症、心房細動、異型狭心症、脳卒中、糖尿病、またはパーキンソン病の診断のための情報提供方法を提供する。
（ａ）正常ヒトおよび被検者のサンプルから分離した小胞からＤＮＡを抽出する段階；
（ｂ）前記抽出したＤＮＡに対して１６Ｓ ｒＤＮＡに存在する遺伝子配列に基づいて作製したプライマーペアを用いてＰＣＲを行って、それぞれのＰＣＲ産物を収得する段階；および
（ｃ）前記ＰＣＲ産物の定量分析を通して正常ヒトに比べてモルガネラ属細菌由来小胞の含量が低い場合、胃癌、大腸癌、すい臓癌、胆管癌、乳癌、卵巣癌、膀胱癌、前立腺癌、リンパ腫、心筋梗塞、心筋症、心房細動、異型狭心症、脳卒中、糖尿病、またはパーキンソン病と判定する段階。 Thus, the present invention comprises the following stages: gastric cancer, colon cancer, pancreatic cancer, bile duct cancer, breast cancer, ovarian cancer, bladder cancer, prostate cancer, lymphoma, myocardial infarction, myocardial disease, atrial fibrillation, atypical angina. Provide informational methods for diagnosing illness, stroke, diabetes, or Parkinson's disease.
(A) Step of extracting DNA from vesicles isolated from normal human and subject samples;
(B) Steps of obtaining each PCR product by performing PCR on the extracted DNA using a primer pair prepared based on the gene sequence present in 16S rDNA; and (c) Quantification of the PCR product. When the content of vesicles derived from Morganella spp. The stage of determining fibrillation, atypical angina, stroke, diabetes, or Parkinson's disease.

本発明において使用される用語「診断」とは、広い意味では患者の病の実態をすべての面にわたって判断することを意味する。判断の内容は、病名、病因、病型、軽重、病床の詳細な様態、合併症の有無、および予後などである。本発明において診断は、胃癌、大腸癌、すい臓癌、胆管癌、乳癌、卵巣癌、膀胱癌、前立腺癌、リンパ腫、心筋梗塞、心筋症、心房細動、異型狭心症、脳卒中、糖尿病、およびパーキンソン病の発病の有無および疾患の水準などを判断することである。 The term "diagnosis" as used in the present invention means to judge the actual condition of a patient's disease in all aspects in a broad sense. The contents of the judgment include the name of the disease, the etiology, the type of the disease, the severity of the disease, the detailed condition of the bed, the presence or absence of complications, and the prognosis. In the present invention, the diagnosis is gastric cancer, colon cancer, pancreatic cancer, bile duct cancer, breast cancer, ovarian cancer, bladder cancer, prostate cancer, lymphoma, myocardial infarction, myocardial infarction, atrial fibrillation, atypical angina, stroke, diabetes, and. It is to judge the presence or absence of Parkinson's disease and the level of the disease.

本発明において使用される用語「ナノ小胞（Ｎａｎｏｖｅｓｉｃｌｅ）あるいは小胞（Ｖｅｓｉｃｌｅ）」とは、多様な細菌から分泌されるナノサイズの膜からなる構造物を意味する。グラム陰性菌（ｇｒａｍ－ｎｅｇａｔｉｖｅ ｂａｃｔｅｒｉａ）由来小胞、または外膜小胞（ｏｕｔｅｒ ｍｅｍｂｒａｎｅ ｖｅｓｉｃｌｅｓ，ＯＭＶｓ）は、内毒素（ｌｉｐｏｐｏｌｙｓａｃｃｈａｒｉｄｅ）だけでなく、毒性タンパク質および細菌ＤＮＡとＲＮＡも有しており、グラム陽性菌（ｇｒａｍ－ｐｏｓｉｔｉｖｅ ｂａｃｔｅｒｉａ）由来小胞は、タンパク質と核酸の他にも、細菌の細胞壁構成成分であるペプチドグリカン（ｐｅｐｔｉｄｏｇｌｙｃａｎ）とリポタイコ酸（ｌｉｐｏｔｅｉｃｈｏｉｃ ａｃｉｄ）も有している。本発明において、ナノ小胞あるいは小胞は、モルガネラ属細菌から自然的に分泌されたり、または人工的に生産されるものであって、球形の形態であり、１０～２００ｎｍの平均直径を有している。 As used in the present invention, the term "Nanovesicle" or "Vesicle" means a structure consisting of nano-sized membranes secreted by various bacteria. Gram-negative bacteria-derived vesicles or outer membrane vesicles (OMVs) have not only endotoxins (lipopolysaccharides), but also toxic proteins and bacterial DNA and RNA, and are gram-positive. In addition to proteins and nucleic acids, gram-positive bacteria-derived follicles also have peptidoglycan and lipotechic acid, which are constituents of the bacterial cell wall. In the present invention, nanovesicles or vesicles are naturally secreted or artificially produced by Morganella spp., Are spherical in shape and have an average diameter of 10-200 nm. ing.

前記小胞は、モルガネラ属細菌を含む培養液を遠心分離、超高速遠心分離、高圧処理、押出、超音波分解、細胞溶解、均質化、冷凍－解凍、電気穿孔、機械的分解、化学物質処理、フィルターによる濾過、ゲル濾過クロマトグラフィー、フリーフロー電気泳動、およびキャピラリー電気泳動よりなる群から選ばれる１つ以上の方法を使用して分離することができる。また、不純物の除去のための洗浄、収得された小胞の濃縮などの過程をさらに含むことができる。 The vesicles are obtained by centrifuging the culture medium containing Morganella spp., Ultrafast centrifugation, high-pressure treatment, extrusion, ultrasonic decomposition, cell lysis, homogenization, freezing-thawing, electroperforation, mechanical decomposition, and chemical treatment. Separation can be performed using one or more methods selected from the group consisting of filtration through a filter, gel filtration chromatography, free-flow electrophoresis, and capillary electrophoresis. Further, processes such as washing for removing impurities and concentration of obtained vesicles can be further included.

本発明において使用される用語「メタゲノム（ｍｅｔａｇｅｎｏｍｅ）」とは、「群 遺伝子」とも言い、土、動物の腸など孤立した地域内のすべてのウイルス、細菌、かびなどを含む遺伝子の総和を意味するものであって、主に培養にならない微生物を分析するために、配列分析器を使用して一度に多くの微生物を同定することを説明するゲノムの概念に使用される。特に、メタゲノムは、一種のゲノムまたは遺伝子を言うものではなく、１つの環境単位のすべての種の遺伝子であって、一種の混合遺伝子を言う。これは、オミックス的に生物学が発展する過程で一種を定義するとき、機能的に既存の一種だけでなく、多様な種が互いに相互作用して完全な種を作るという観点から出た用語である。技術的には、速い配列分析法を利用して、種に関係なく、すべてのＤＮＡ、ＲＮＡを分析して、１つの環境内でのすべての種を同定し、相互作用、代謝作用を解明する技法の対象である。 The term "metagenomic" used in the present invention is also referred to as "group gene" and means the sum of genes including all viruses, bacteria, molds, etc. in isolated areas such as soil and animal intestines. It is mainly used in the concept of genome to explain the identification of many microorganisms at once using a sequence analyzer to analyze non-cultivated microorganisms. In particular, metagenomics does not refer to a kind of genome or gene, but to all kinds of genes in one environmental unit, and refers to a kind of mixed gene. This is a term that comes from the perspective that when defining a species in the process of omics-like development of biology, not only functionally existing species but also diverse species interact with each other to form a complete species. be. Technically, a fast sequence analysis method is used to analyze all DNA and RNA regardless of species, identify all species in one environment, and elucidate interactions and metabolic effects. It is the subject of the technique.

本発明において、前記段階（ａ）でのサンプルは、血液、尿、または便でありうるが、これに制限されるものではない。 In the present invention, the sample in step (a) can be blood, urine, or stool, but is not limited thereto.

本発明において、前記段階（ｂ）でのプライマーペアは、配列番号１および配列番号２のプライマーでありうる。 In the present invention, the primer pair in the step (b) can be the primers of SEQ ID NO: 1 and SEQ ID NO: 2.

本発明の他の様態として、本発明は、モルガネラ属細菌由来小胞を有効成分として含む、胃癌、大腸癌、すい臓癌、胆管癌、乳癌、卵巣癌、膀胱癌、前立腺癌、リンパ腫、心筋梗塞、心筋症、心房細動、異型狭心症、脳卒中、糖尿病、パーキンソン病、および炎症性疾患よりなる群から選ばれる１つ以上の疾病の予防または治療用薬学的組成物を提供する。 As another aspect of the present invention, the present invention contains gastric cancer, colon cancer, pancreatic cancer, bile duct cancer, breast cancer, ovarian cancer, bladder cancer, prostate cancer, lymphoma, myocardial infarction, which contain vesicles derived from Morganella spp. , Atypical angina, stroke, diabetes, Parkinson's disease, and a pharmaceutical composition for the prevention or treatment of one or more diseases selected from the group consisting of inflammatory diseases.

本発明のさらに他の様態として、本発明は、モルガネラ属細菌由来小胞を有効成分として含む、胃癌、大腸癌、すい臓癌、胆管癌、乳癌、卵巣癌、膀胱癌、前立腺癌、リンパ腫、心筋梗塞、心筋症、心房細動、異型狭心症、脳卒中、糖尿病、パーキンソン病、および炎症性疾患よりなる群から選ばれる１つ以上の疾病の予防または改善用食品組成物を提供する。 As yet another aspect of the present invention, the present invention contains gastric cancer, colon cancer, pancreatic cancer, bile duct cancer, breast cancer, ovarian cancer, bladder cancer, prostate cancer, lymphoma, myocardial infarction containing vesicles derived from Morganella spp. Provided are food compositions for the prevention or amelioration of one or more diseases selected from the group consisting of infarction, myocardial infarction, atrial fibrillation, atypical angina, stroke, diabetes, Parkinson's disease, and inflammatory disease.

本発明において、前記炎症性疾患は、アトピー皮膚炎、にきび、乾癬、副鼻腔炎、鼻炎、結膜炎、喘息、皮膚炎、炎症性コラーゲン血管疾患、糸球体腎炎、脳炎、炎症性腸炎、慢性閉鎖性肺疾患、敗血症、敗血症性ショック、肺線維症、未分化脊椎関節症、未分化関節症、関節炎、炎症性骨溶解、ウイルスまたはバクテリア感染による慢性炎症疾患、大腸炎、潰瘍性大腸炎、炎症性腸疾患、関節炎、関節リウマチ、反応性関節炎、骨関節炎、鞏皮症、骨粗しょう症、アテローム性動脈硬化症、心筋炎、心内膜炎、心嚢炎、嚢胞性線維症、橋本甲状腺炎、グレーブス病、ハンセン病、梅毒、ライム病（Ｌｙｍｅ ｄｉｓｅａｓｅ）、ボレリア症（Ｂｏｒｒｅｌｉｏｓｉｓ）、神経性ボレリア症、結核、サルコイドーシス（Ｓａｒｃｏｉｄｏｓｉｓ）、ループス、凍瘡状ループス、結核性ループス、ループス腎炎、全身性エリテマトーデス、黄斑変性、ブドウ膜炎、過敏性腸症候群、クローン病、シェーグレン症候群、線維筋痛、慢性疲労症候群、慢性疲労免疫不全症候群、筋痛性脳脊髄炎、筋萎縮性側索硬化症、パーキンソン病、および多発性硬化症よりなる群から選ばれる１つ以上でありうるが、これに制限されない。 In the present invention, the inflammatory disease includes atopy dermatitis, acne, psoriasis, sinusitis, rhinitis, conjunctivitis, asthma, dermatitis, inflammatory collagen vascular disease, glomerular nephritis, encephalitis, inflammatory enteritis, chronic closure. Pulmonary disease, septicemia, septic shock, pulmonary fibrosis, undifferentiated lupus erythematosus, undifferentiated arthritis, arthritis, inflammatory osteolysis, chronic inflammatory disease due to viral or bacterial infection, colitis, ulcerative colitis, inflammatory Intestinal disease, arthritis, rheumatoid arthritis, reactive arthritis, osteoarthritis, erythematosus, osteoporosis, atherosclerosis, myocarditis, endocarditis, cystitis, cystic fibrosis, Hashimoto thyroiditis, Lyme disease Diseases, Hansen's disease, syphilis, Lyme disease, Borreliosis, neuroborreliosis, tuberculosis, sarcoidosis, lupus, frost-like lupus, tuberculous lupus, lupus erythematosus, lupus erythematosus, lupus erythematosus , Lyme disease, irritable intestinal syndrome, Crohn's disease, Schegren's syndrome, fibromyalgia, chronic fatigue syndrome, chronic fatigue immunodeficiency syndrome, myopathic encephalomyelitis, my atrophic lateral sclerosis, Parkinson's disease, and multiple It can be, but is not limited to, one or more selected from the group consisting of sclerosis.

本発明において、前記炎症性疾患は、インターロイキン－６（Ｉｎｔｅｒｌｅｕｋｉｎ－６；ＩＬ－６）または腫瘍壊死因子－アルファ（Ｔｕｍｏｒ ｎｅｃｒｏｓｉｓ ｆａｃｔｏｒ－ａｌｐｈａ；ＴＮＦ－α）により媒介される疾患でありうるが、これに制限されない。 In the present invention, the inflammatory disease can be a disease mediated by interleukin-6 (Interleukin-6; IL-6) or tumor necrosis factor-alpha (TNF-α). Not limited to this.

本発明において使用される用語「予防」とは、本発明による食品または薬学的組成物の投与により癌、炎症疾患、心血管疾患、代謝疾患、または神経－精神疾患を抑制させたり発病を遅延させるすべての行為を意味する。 The term "prevention" as used in the present invention means that administration of a food or pharmaceutical composition according to the present invention suppresses or delays the onset of cancer, inflammatory disease, cardiovascular disease, metabolic disease, or neuro-psychiatric disease. Means all actions.

本発明において使用される用語「治療」とは、本発明による薬学的組成物の投与により癌、炎症疾患、心血管疾患、代謝疾患、または神経－精神疾患に対する症状が好転したり有利に変更されるすべての行為を意味する。 The term "treatment" as used in the present invention means that administration of the pharmaceutical composition according to the present invention improves or favorably changes symptoms for cancer, inflammatory disease, cardiovascular disease, metabolic disease, or neuro-psychiatric disease. Means all actions.

本発明において使用される用語「改善」とは、治療される状態と関連したパラメーター、例えば症状の程度を少なくとも減少させるすべての行為を意味する。 As used in the present invention, the term "improvement" means any action that at least reduces the degree of a condition associated with the condition being treated, eg, the degree of symptom.

本発明の一実施例では、細菌および細菌由来小胞をマウスの経口で投与して細菌および小胞の体内吸収、分布、および排泄様相を評価し、細菌である場合には、腸粘膜を通じて吸収されないのに対して、小胞は、投与５分以内に吸収されて全身的に分布し、腎臓、肝臓などを通じて排泄されることを確認した（実施例１参照）。 In one embodiment of the invention, bacteria and vesicles derived from the bacterium are administered orally in mice to evaluate the absorption, distribution, and excretory aspects of the bacteria and vesicles, and if they are bacteria, absorbed through the intestinal mucosa. On the other hand, it was confirmed that the vesicles were absorbed within 5 minutes after administration, distributed systemically, and excreted through the kidney, liver, etc. (see Example 1).

本発明の他の一実施例では、胃癌、大腸癌、すい臓癌、胆管癌、乳癌、卵巣癌、膀胱癌、前立腺癌、リンパ腫、心筋梗塞、心筋症、心房細動、異型狭心症、脳卒中、糖尿病、およびパーキンソン病の患者に年齢と性別をマッチングした正常ヒトの血液、尿、または便から分離した小胞を用いて細菌メタゲノム分析を実施した。その結果、正常ヒトのサンプルに比べて、胃癌、大腸癌、すい臓癌、胆管癌、乳癌、卵巣癌、膀胱癌、前立腺癌、リンパ腫、心筋梗塞、心筋症、心房細動、異型狭心症、脳卒中、糖尿病、およびパーキンソン病の患者のサンプルにモルガネラ属細菌由来小胞が有意に減少していることを確認した（実施例３～１５参照）。 In another embodiment of the invention, gastric cancer, colon cancer, pancreatic cancer, bile duct cancer, breast cancer, ovarian cancer, bladder cancer, prostate cancer, lymphoma, myocardial infarction, myocardial disease, atrial fibrillation, atypical angina, stroke. Bacterial metagenome analysis was performed on patients with diabetes, diabetes, and Parkinson's disease using vesicles isolated from normal human blood, urine, or stool that matched age and gender. As a result, compared to normal human samples, gastric cancer, colon cancer, pancreatic cancer, bile duct cancer, breast cancer, ovarian cancer, bladder cancer, prostate cancer, lymphoma, myocardial infarction, myocardial disease, atrial fibrillation, atypical angina, It was confirmed that the follicles derived from Morganella spp. Were significantly reduced in the samples of patients with stroke, diabetes, and Parkinson's disease (see Examples 3 to 15).

本発明のさらに他の実施例では、炎症細胞でモルガネラ・モルガニイ由来小胞（Ｍ．ｍｏｒｇａｎｉｉ ＥＶ）の細胞死滅効果を評価するために、マウスマクロファージ細胞株であるＲａｗ ２６４．７細胞にモルガネラ・モルガニイ（ＭＭＲ１０１、ＭＭＲ２０１、ＭＭＲ２０２）由来小胞を多様な濃度（０．１、１、１０μｇ／ｍｌ）で処理した後、細胞死滅程度を評価した結果、モルガネラ・モルガニイ（ＭＭＲ１０１、ＭＭＲ２０１、ＭＭＲ２０２）由来小胞の処理時に細胞死滅は観察されないことを確認した（実施例１７参照）。 In yet another embodiment of the invention, Morganella morganii was added to Raw 264.7 cells, a mouse macrophage cell line, to evaluate the cell-killing effect of M. morganii EV on inflammatory cells. After treating vesicles derived from (MMR101, MMR201, MMR202) at various concentrations (0.1, 1, 10 μg / ml) and evaluating the degree of cell death, small vesicles derived from Morganella Morganii (MMR101, MMR201, MMR202) It was confirmed that no cell death was observed during vesicle treatment (see Example 17).

本発明のさらに他の実施例では、前記実施例の結果に基づいてモルガネラ属細菌に属するモルガネラ・モルガニイ種細菌由来小胞の特性を分析するためにさらに研究した結果、モルガネラ・モルガニイ菌株を培養してこれから分泌された小胞が抗炎症効果を示すかを評価したが、多様な濃度のモルガネラ・モルガニイ由来小胞をマクロファージに処理した後、炎症疾患の原因因子である大腸菌由来小胞を処理して炎症メディエーターの分泌を評価した結果、大腸菌由来小胞によるＩＬ－６およびＴＮＦ－αの分泌をモルガネラ・モルガニイ由来小胞が効率的に抑制することを確認した（実施例１８参照）。 In still another example of the present invention, as a result of further research to analyze the characteristics of vesicles derived from Morganella Morganella spp. We evaluated whether the vesicles secreted from now on showed an anti-inflammatory effect. After treating macrophages with various concentrations of Morganella-Morganella-derived vesicles, we treated Escherichia coli-derived vesicles, which are the causative factors of inflammatory diseases. As a result of evaluating the secretion of the inflammatory mediator, it was confirmed that the Morganella-Morganella-derived vesicles efficiently suppressed the secretion of IL-6 and TNF-α by the E. coli-derived vesicles (see Example 18).

本発明のさらに他の実施例では、モルガネラ・モルガニイ由来小胞の抗炎症作用が熱処理または酸処理の効果を評価するために、マクロファージに大腸菌由来小胞を処理する前に熱処理または酸処理した小胞を投与してＴＮＦ－αの分泌に及ぼす影響を評価した結果、モルガネラ・モルガニイ由来小胞によるＴＮＦ－αの分泌抑制効果が熱処理や酸処理により変わらないことを確認した（実施例１９参照）。 In yet another embodiment of the invention, the anti-inflammatory effect of Morganella morganii-derived vesicles is heat-treated or acid-treated prior to treating macrophages with E. coli-derived vesicles to assess the effect of heat treatment or acid treatment. As a result of evaluating the effect on the secretion of TNF-α by administering the vesicles, it was confirmed that the effect of suppressing the secretion of TNF-α by the vesicles derived from Morganella morganii did not change by heat treatment or acid treatment (see Example 19). ..

本発明のさらに他の実施例では、モルガネラ・モルガニイ菌株を培養してこれから分泌された小胞が抗癌治療効果を示すかを評価した。このために、癌細胞株を皮下で注射して癌モデルを作成し、モルガネラ・モルガニイ由来小胞を癌細胞株の処置４日前からマウスに経口または腹腔で投与した後、２０日間癌組織の大きさを測定した結果、前記小胞を腹腔および経口で投与した場合に、対照群に比べて癌組織の大きさが減少し、特に経口で投与した場合に顕著に減少していることを確認した（実施例２０参照）。 In still another example of the present invention, the Morganella-Morganella strain was cultured and the vesicles secreted from the vesicles were evaluated to have an anti-cancer therapeutic effect. For this purpose, a cancer cell line is injected subcutaneously to create a cancer model, and vesicles derived from Morganella morganii are orally or peritoneally administered to mice 4 days before treatment of the cancer cell line, and then the size of the cancer tissue is 20 days. As a result of the measurement, it was confirmed that when the vesicles were administered intraperitoneally and orally, the size of the cancer tissue was reduced as compared with the control group, and in particular, when the vesicles were orally administered, the size was significantly reduced. (See Example 20).

本発明による薬学的組成物は、薬学的に許容可能な担体を含むことができる。前記薬学的に許容可能な担体は、製剤時に通常用いられるものであって、食塩水、滅菌水、リンガー液、緩衝食塩水、シクロデキストリン、デキストロース溶液、マルトデキストリン溶液、グリセロール、エタノール、リポソームなどを含むが、これに限定されず、必要に応じて抗酸化剤、緩衝液など他の通常の添加剤をさらに含むことができる。また、希釈剤、分散剤、界面活性剤、結合剤、潤滑剤などを付加的に添加して水溶液、懸濁液、乳濁液などのような注射用剤形、丸薬、カプセル、顆粒、または錠剤に製剤化することができる。適切な薬学的に許容される担体および製剤化に関しては、レミントンの文献に開示されている方法を用いて各成分によって好適に製剤化することができる。本発明の薬学的組成物は、剤形に特別な制限はないが、注射剤、吸入剤、皮膚外用剤、または経口摂取剤などに製剤化することができる。 The pharmaceutical composition according to the invention can include a pharmaceutically acceptable carrier. The pharmaceutically acceptable carrier is usually used at the time of preparation, and includes saline solution, sterile water, Ringer's solution, buffered saline solution, cyclodextrin, dextrose solution, maltodextrin solution, glycerol, ethanol, liposomes and the like. Including, but not limited to, other usual additives such as antioxidants, buffers and the like can be further included as needed. In addition, additional additives such as diluents, dispersants, surfactants, binders, lubricants, etc. are added to form injectable forms such as aqueous solutions, suspensions, emulsions, pills, capsules, granules, or It can be formulated into tablets. For appropriate pharmaceutically acceptable carriers and formulations, each component can be suitably formulated using the methods disclosed in Remington's literature. The pharmaceutical composition of the present invention can be formulated into an injection, an inhalant, an external skin preparation, an oral ingestion, or the like, although the dosage form is not particularly limited.

本発明の薬学的組成物は、目的とする方法によって経口投与したり、非経口投与（例えば、静脈内、皮下、皮膚、鼻腔、気道に適用）することができ、投与量は、患者の状態および体重、疾病の程度、薬物形態、投与経路および時間によって異なるが、当業者により適宜選択され得る。 The pharmaceutical composition of the present invention can be orally administered or parenterally (eg, applied intravenously, subcutaneously, skin, nasal cavity, airway) by a method of interest, and the dose is the patient's condition. And depending on body weight, degree of disease, drug form, route of administration and time, but may be appropriately selected by those skilled in the art.

本発明による薬学的組成物は、薬学的に有効な量で投与する。本発明において、薬学的に有効な量は、医学的治療に適用可能な合理的な利益／リスクの割合で疾患を治療するのに十分な量を意味し、有効用量の水準は、患者の疾患の種類、重症度、薬物の活性、薬物に対する敏感度、投与時間、投与経路および排出比率、治療期間、同時使用される薬物を含む要素およびその他医学分野によく知られた要素によって決定され得る。本発明による組成物は、個別治療剤として投与したり他の治療剤と併用して投与することができ、従来の治療剤とは順次または同時に投与され得、単一または多重投与され得る。上記した要素を全部考慮して副作用なしに最小の量で最大の効果を得ることができる量を投与することが重要であり、これは、当業者によって容易に決定され得る。 The pharmaceutical composition according to the invention is administered in a pharmaceutically effective amount. In the present invention, a pharmaceutically effective amount means an amount sufficient to treat a disease at a reasonable benefit / risk ratio applicable to medical treatment, and an effective dose level is a patient's disease. It can be determined by the type, severity, activity of the drug, sensitivity to the drug, duration of administration, route and excretion ratio, duration of treatment, factors including concomitant drugs and other factors well known in the medical field. The compositions according to the invention can be administered as individual therapeutic agents or in combination with other therapeutic agents, can be administered sequentially or simultaneously with conventional therapeutic agents, and can be administered single or multiple times. It is important to consider all of the above factors and administer an amount that will give the maximum effect with the minimum amount without side effects, which can be easily determined by those skilled in the art.

具体的に、本発明による薬学的組成物の有効量は、患者の年齢、性別、体重によって変わり得、一般的には、体重１ｋｇ当たり０．００１～１５０ｍｇ、好ましくは０．０１～１００ｍｇを毎日または隔日投与したり、１日１～３回に分けて投与することができる。しかしながら、投与経路、肥満の重症度、性別、体重、年齢などによって増減され得るので、前記投与量がいかなる意味でも本発明の範囲を限定するものではない。 Specifically, the effective amount of the pharmaceutical composition according to the invention may vary depending on the age, gender and body weight of the patient, generally 0.001 to 150 mg / kg body weight, preferably 0.01 to 100 mg daily. Alternatively, it can be administered every other day or divided into 1 to 3 times a day. However, since the dose may be increased or decreased depending on the route of administration, the severity of obesity, gender, body weight, age, etc., the dose does not limit the scope of the present invention in any sense.

本発明の食品組成物は、健康機能食品組成物を含む。本発明による食品組成物は、有効成分を食品にそのまま添加したり他の食品または食品成分と共に使用され得、通常の方法によって適切に使用され得る。有効成分の混合量は、その使用目的（予防または改善用）によって適宜決定され得る。一般的に、食品または飲料の製造時に、本発明の組成物は、原料に対して１５重量％以下、好ましくは１０重量％以下の量で添加される。しかしながら、健康および衛生を目的としたりまたは健康調節を目的とする長期間の摂取の場合には、前記量は、前記範囲以下でありうる。 The food composition of the present invention includes a health functional food composition. The food composition according to the present invention can be added as it is to a food or used in combination with other foods or food ingredients, and can be appropriately used by a usual method. The mixing amount of the active ingredient may be appropriately determined depending on the purpose of use (prevention or improvement). Generally, during the production of foods or beverages, the compositions of the present invention are added in an amount of 15% by weight or less, preferably 10% by weight or less, based on the raw materials. However, in the case of long-term ingestion for the purpose of health and hygiene or for the purpose of health regulation, the amount may be less than or equal to the above range.

本発明の食品組成物は、指示された割合で必須成分として前記有効成分を含有すること他に、他の成分には特別な制限がなく、通常の飲料のように様々な香味剤または天然炭水化物などをさらなる成分として含有することができる。上述した天然炭水化物の例は、単糖類、例えば、ブドウ糖、果糖など；二糖類、例えばマルトース、スクロースなど；および多糖類、例えばデキストリン、シクロデキストリンなどのような通常の糖、およびキシリトール、ソルビトール、エリスリトールなどの糖アルコールである。上述したもの以外の香味剤として天然香味剤（ソーマチン、ステビア抽出物（例えばレバウジオシドＡ、グリシルリジンなど）および合成香味剤（サッカリン、アスパルテームなど）を有利に使用することができる。前記天然炭水化物の比率は、当業者の選択によって適切に決定され得る。 In addition to containing the active ingredient as an essential ingredient in the specified proportion, the food composition of the present invention has no special restrictions on other ingredients, and various flavoring agents or natural carbohydrates like ordinary beverages. Etc. can be contained as a further component. Examples of natural carbohydrates mentioned above include monosaccharides such as glucose, fructose; disaccharides such as maltose, sucrose, etc .; and polysaccharides such as common sugars such as dextrin, cyclodextrin, etc., and xylitol, sorbitol, erythritol, etc. Such as sugar alcohol. Natural flavoring agents (such as thaumatin, stevia extract (eg, levaugioside A, glycyrrhizin, etc.) and synthetic flavoring agents (saccharin, aspartame, etc.) can be advantageously used as flavoring agents other than those described above. , Can be appropriately determined by the choice of one of ordinary skill in the art.

前記の他に本発明の食品組成物は、様々な栄養剤、ビタミン、ミネラル（電解質）、合成風味剤および天然風味剤などの風味剤、着色剤および充填剤（チーズ、チョコレートなど）、ペクチン酸およびその塩、アルギン酸およびその塩、有機酸、保護性コロイド増粘剤、ｐＨ調節剤、安定化剤、防腐剤、グリセリン、アルコール、炭酸飲料に使用される炭酸化剤などを含有することができる。このような成分は、独立して、または組み合わせて使用することができる。このような添加剤の比率も、当業者によって適宜選択され得る。 In addition to the above, the food compositions of the present invention include various nutrients, vitamins, minerals (electrolytes), flavoring agents such as synthetic flavoring agents and natural flavoring agents, coloring agents and fillers (cheese, chocolate, etc.), pectic acids. And its salts, alginic acid and its salts, organic acids, protective colloid thickeners, pH regulators, stabilizers, preservatives, glycerin, alcohols, carbonizing agents used in carbonated beverages and the like. .. Such ingredients can be used independently or in combination. The ratio of such additives may also be appropriately selected by those skilled in the art.

以下、本発明の理解を助けるために好適な実施例を提示する。しかしながら、下記の実施例は、本発明をさらに容易に理解するために提供されるものに過ぎず、下記実施例によって本発明の内容が制限されるものではない。 Hereinafter, suitable examples will be presented in order to help the understanding of the present invention. However, the following examples are provided only for easier understanding of the present invention, and the contents of the present invention are not limited by the following examples.

［実施例１．腸内細菌および細菌由来小胞の体内吸収、分布、および排泄様相の分析］
腸内細菌と細菌由来小胞が胃腸管を通じて全身的に吸収されるかを評価するために、次のような方法で実験を行った。マウスの胃腸に蛍光で標識した腸内細菌と腸内細菌由来小胞をそれぞれ５０μｇの用量で胃腸管に投与し、０分、５分、３時間、６時間、１２時間後に蛍光を測定した。マウス全体像を観察した結果、図１ａに示されたように、細菌である場合には、腸粘膜を通じて全身的に吸収されないが、細菌由来小胞である場合には、投与後５分に全身的に吸収され、投与３０分には、膀胱に蛍光が濃く観察されて、小胞が泌尿器系に排泄されることが分かった。また、小胞は、投与１２時間まで体内に存在することが分かった（図１ａ参照）。 [Example 1. Analysis of absorption, distribution, and excretion aspects of gut microbiota and bacterial vesicles]
In order to evaluate whether intestinal bacteria and bacterial vesicles are systemically absorbed through the gastrointestinal tract, experiments were performed by the following methods. Gut microbiota labeled with fluorescence in the gastrointestinal tract of mice and vesicles derived from the gut microbiota were administered to the gastrointestinal tract at doses of 50 μg, respectively, and fluorescence was measured after 0 minutes, 5 minutes, 3 hours, 6 hours, and 12 hours. As a result of observing the whole image of the mouse, as shown in FIG. 1a, in the case of bacteria, it is not systemically absorbed through the intestinal mucosa, but in the case of bacterial vesicles, it is systemically absorbed 5 minutes after administration. At 30 minutes after administration, a strong fluorescence was observed in the bladder, indicating that vesicles were excreted in the urinary system. It was also found that vesicles were present in the body up to 12 hours after administration (see FIG. 1a).

腸内細菌と腸内細菌由来小胞が全身的に吸収された後、様々な臓器に浸潤された様相を評価するために、蛍光で標識した５０μｇの細菌と細菌由来小胞を前記の方法のように投与した後、投与１２時間後に血液、心臓、肝臓、腎臓、脾臓、脂肪、筋肉を採取した。採取した組織から蛍光を観察した結果、図１ｂに示されたように、細菌由来小胞が、血液、心臓、肺、肝臓、腎臓、脾臓、脂肪、筋肉、腎臓に分布したが、細菌は吸収されないことが分かった（図１ｂ参照）。 After systemic absorption of Gut microbiota and Gut microbiota-derived vesicles, 50 μg of fluorescently labeled bacteria and bacterial-derived vesicles were added to the method described above to assess the appearance of infiltration into various organs. Blood, heart, liver, kidney, spleen, fat, and muscle were collected 12 hours after administration. As a result of observing fluorescence from the collected tissue, as shown in FIG. 1b, bacterial-derived vesicles were distributed in blood, heart, lung, liver, kidney, spleen, fat, muscle, and kidney, but the bacteria were absorbed. It turned out not to be done (see Figure 1b).

［実施例２．臨床サンプルで細菌由来小胞メタゲノム分析］
血液、尿、便などの臨床サンプルをまず１０ｍｌチューブに入れて遠心分離法（３，５００ｘｇ、１０ｍｉｎ、４℃）で浮遊物を沈め、上澄み液だけを新しい１０ｍｌチューブに移した。０．２２μｍのフィルターを使用して細菌および異物を除去した後、セントリプレップチューブ（Ｃｅｎｔｒｉｐｒｅｐ ｃｅｎｔｒｉｆｕｇａｌ ｆｉｌｔｅｒｓ ５０ｋＤ）に移して１５００×ｇ、４℃で１５分間遠心分離して、５０ｋＤより小さい物質は捨てて、１０ｍｌまで濃縮させた。さらに０．２２μｍのフィルターを使用してバクテリアおよび異物を除去した後、Ｔｙｐｅ ９０ｔｉローターで１５０，０００×ｇ、４℃で３時間の間超高速遠心分離方法を使用して上澄み液を捨てて、固まったペレットを生理食塩水（ＰＢＳ）で溶かした。 [Example 2. Bacterial vesicle metagenomic analysis in clinical samples]
Clinical samples such as blood, urine, and stool were first placed in a 10 ml tube, the suspension was submerged by centrifugation (3,500 xg, 10 min, 4 ° C.), and only the supernatant was transferred to a new 10 ml tube. After removing bacteria and foreign matter using a 0.22 μm filter, transfer to a Centriprep tube (Centriprep centrifugal filters 50 kD), centrifuge at 1500 × g at 4 ° C for 15 minutes, and discard substances smaller than 50 kD. It was concentrated to 10 ml. After further removing bacteria and foreign matter using a 0.22 μm filter, discard the supernatant using an ultrafast centrifugation method at 150,000 xg, 4 ° C. for 3 hours on a Type 90ti rotor. The solidified pellet was dissolved in saline (PBS).

前記方法で分離した小胞１００μｌを１００℃で沸かして内部のＤＮＡを脂質外に出るようにし、その後、氷上で５分間冷ました。そして、残った浮遊物を除去するために、１０，０００×ｇ、４℃で３０分間遠心分離し、上澄み液だけを集めた。そして、Ｎａｎｏｄｒｏｐを用いてＤＮＡ量を定量した。その後、前記抽出されたＤＮＡに細菌由来ＤＮＡが存在するかを確認するために、下記表１に示した１６ｓ ｒＤＮＡ ｐｒｉｍｅｒでＰＣＲを行って、前記抽出された遺伝子に細菌由来遺伝子が存在することを確認した。 100 μl of the vesicles separated by the above method were boiled at 100 ° C. to allow the internal DNA to come out of the lipid, and then cooled on ice for 5 minutes. Then, in order to remove the remaining suspended matter, the mixture was centrifuged at 10,000 × g at 4 ° C. for 30 minutes, and only the supernatant liquid was collected. Then, the amount of DNA was quantified using Nanodrop. Then, in order to confirm the presence of bacterial-derived DNA in the extracted DNA, PCR was performed with the 16s rDNA primer shown in Table 1 below to confirm that the extracted gene had a bacterial-derived gene. confirmed.

Figure 2022028794000002
Figure 2022028794000002

前記方法で抽出したＤＮＡを前記の１６Ｓ ｒＤＮＡプライマーを使用して増幅した後、シーケンシングを行い（Ｉｌｌｕｍｉｎａ ＭｉＳｅｑ ｓｅｑｕｅｎｃｅｒ）、その結果をＳｔａｎｄａｒｄ Ｆｌｏｗｇｒａｍ Ｆｏｒｍａｔ（ＳＦＦ）ファイルで出力してＧＳ ＦＬＸ ｓｏｆｔｗａｒｅ（ｖ２．９）を利用してＳＦＦファイルをｓｅｑｕｅｎｃｅファイル（．ｆａｓｔａ）とｎｕｃｌｅｏｔｉｄｅ ｑｕａｌｉｔｙ ｓｃｏｒｅファイルに変換した後、リードの信用度評価を確認し、ｗｉｎｄｏｗ（２０ｂｐｓ）平均ｂａｓｅ ｃａｌｌ ａｃｃｕｒａｃｙが９９％未満（Ｐｈｒｅｄ ｓｃｏｒｅ＜２０）である部分を除去した。Ｏｐｅｒａｔｉｏｎａｌ Ｔａｘｏｎｏｍｙ Ｕｎｉｔ（ＯＴＵ）分析のためには、ＵＣＬＵＳＴとＵＳＥＡＲＣＨを利用してシークエンス類似度によってクラスタリングを行い、ｇｅｎｕｓは９４％、ｆａｍｉｌｙは９０％、ｏｒｄｅｒは８５％、ｃｌａｓｓは８０％、ｐｈｙｌｕｍは７５％シークエンス類似度を基準としてクラスタリングを行い、各ＯＴＵのｐｈｙｌｕｍ、ｃｌａｓｓ、ｏｒｄｅｒ、ｆａｍｉｌｙ、ｇｅｎｕｓレベルの分類を行い、ＢＬＡＳＴＮとＧｒｅｅｎＧｅｎｅｓの１６Ｓ ＲＮＡシークエンスデータベース（１０８，４５３シークエンス）を用いて属水準で９７％以上のシークエンス類似度を有する細菌をプロファイリングした（ＱＩＩＭＥ）。 The DNA extracted by the above method is amplified using the above 16S rDNA primer, then sequenced (Illumina MiSeq sequencer), and the result is output as a Standard Flowgram Format (SFF) file to output the GS FLX software (v2.). After converting the SFF file into a sequence file (.fasta) and a nucleotide quality score file using 9), the credit rating of the lead is confirmed, and the window (20 bps) average base call currency is less than 99% (Phred score <20). ) Was removed. For Operational Taxonomy Unit (OTU) analysis, clustering is performed by sequence similarity using UCLUST and USEARCH, genus is 94%, family is 90%, orderer is 85%, class is 80%, and phylum is 75. % Sequence Similarity is used for clustering, each OTU is classified into phylum, class, orderer, family, and genus levels, and BLASTN and GreenGenes 16S RNA sequence database (108,453 sequences) is used to classify 97 at the genus level. Bacteria with a sequence similarity of% or higher were profiled (QIIME).

［実施例３．胃癌患者の便、血液および尿細菌由来小胞メタゲノム分析］
実施例２の方法で胃癌患者６３人および、年齢と性別をマッチングした正常ヒト１２６人の便を対象に、便内に存在する小胞から遺伝子を抽出してメタゲノム分析を行った後、モルガネラ属細菌由来小胞の分布を評価した。その結果、正常ヒトの便に比べて胃癌患者の便にモルガネラ属細菌由来小胞が有意に減少していることを確認した（表２および図２ａ参照）。 [Example 3. Metagenomic analysis of vesicles derived from stool, blood and urinary bacteria of gastric cancer patients]
In the stools of 63 gastric cancer patients and 126 normal humans whose age and gender were matched by the method of Example 2, genes were extracted from vesicles present in the stools and metagenome analysis was performed, and then Morganella spp. The distribution of bacterial vesicles was evaluated. As a result, it was confirmed that vesicles derived from Morganella spp. Were significantly reduced in the stools of gastric cancer patients as compared with the stools of normal humans (see Table 2 and FIG. 2a).

Figure 2022028794000003
Figure 2022028794000003

実施例２の方法で胃癌患者６６人および、年齢と性別をマッチングした正常ヒト１９８人の血液を対象に、血液内に存在する小胞から遺伝子を抽出してメタゲノム分析を行った後、モルガネラ属細菌由来小胞の分布を評価した。その結果、正常ヒトの血液に比べて胃癌患者の血液にモルガネラ属細菌由来小胞が有意に減少していることを確認した（表３および図２ｂ参照）。 In the blood of 66 gastric cancer patients and 198 normal humans whose age and gender were matched by the method of Example 2, genes were extracted from vesicles present in the blood and metagenome analysis was performed, and then Morganella spp. The distribution of bacterial vesicles was evaluated. As a result, it was confirmed that vesicles derived from Morganella spp. Were significantly reduced in the blood of gastric cancer patients as compared with the blood of normal humans (see Table 3 and FIG. 2b).

Figure 2022028794000004
Figure 2022028794000004

また、実施例２の方法で胃癌患者６１人の尿および、性別と年齢をマッチングした正常対照群１２０人の尿を対象に、尿内に存在する小胞から遺伝子を抽出してメタゲノム分析を行った後、モルガネラ属細菌由来小胞の分布を評価した。その結果、正常ヒトの尿に比べて胃癌患者の尿にモルガネラ属細菌由来小胞が有意に減少していることを確認した（表４および図２ｃ参照）。 In addition, by the method of Example 2, genes were extracted from vesicles present in urine and metagenome analysis was performed on the urine of 61 gastric cancer patients and the urine of 120 normal control groups whose sex and age were matched. After that, the distribution of vesicles derived from Morganella spp. Was evaluated. As a result, it was confirmed that vesicles derived from Morganella spp. Were significantly reduced in the urine of gastric cancer patients as compared with the urine of normal humans (see Table 4 and FIG. 2c).

Figure 2022028794000005
Figure 2022028794000005

［実施例４．大腸癌患者の尿細菌由来小胞メタゲノム分析］
実施例２の方法で大腸癌患者３８人および、年齢と性別をマッチングした正常ヒト３８人の尿を対象に、尿内に存在する小胞から遺伝子を抽出してメタゲノム分析を行った後、モルガネラ属細菌由来小胞の分布を評価した。その結果、正常ヒトの尿に比べて大腸癌患者の尿にモルガネラ属細菌由来小胞が有意に減少していることを確認した（表５および図３参照）。 [Example 4. Metagenomic analysis of vesicles derived from urinary bacteria in colorectal cancer patients]
In the urine of 38 colorectal cancer patients and 38 normal humans whose age and gender were matched by the method of Example 2, genes were extracted from vesicles present in the urine and metagenome analysis was performed, and then Morganella was performed. The distribution of vesicles derived from the genus bacteria was evaluated. As a result, it was confirmed that vesicles derived from Morganella spp. Are significantly reduced in the urine of colorectal cancer patients as compared with the urine of normal humans (see Table 5 and FIG. 3).

Figure 2022028794000006
Figure 2022028794000006

［実施例５．すい臓癌患者の血液細菌由来小胞メタゲノム分析］
実施例２の方法ですい臓癌患者１７６人および、年齢と性別をマッチングした正常ヒト２７１人の血液を対象に、血液内に存在する小胞から遺伝子を抽出してメタゲノム分析を行った後、モルガネラ属細菌由来小胞の分布を評価した。その結果、正常ヒトの血液に比べてすい臓癌患者の血液にモルガネラ属細菌由来小胞が有意に減少していることを確認した（表６および図４参照）。 [Example 5. Metagenomic analysis of vesicles derived from blood bacteria in patients with pancreatic cancer]
In the blood of 176 patients with pancreatic cancer and 271 normal humans whose age and gender were matched by the method of Example 2, genes were extracted from vesicles existing in the blood and metagenome analysis was performed, and then Morganella was performed. The distribution of vesicles derived from the genus bacteria was evaluated. As a result, it was confirmed that vesicles derived from Morganella spp. Are significantly reduced in the blood of pancreatic cancer patients as compared with the blood of normal humans (see Table 6 and FIG. 4).

Figure 2022028794000007
Figure 2022028794000007

［実施例６．胆管癌患者の血液細菌由来小胞メタゲノム分析］
実施例２の方法で胆管癌患者７９人および、年齢と性別をマッチングした正常ヒト２５９人の血液を対象に、血液内に存在する小胞から遺伝子を抽出してメタゲノム分析を行った後、モルガネラ属細菌由来小胞の分布を評価した。その結果、正常ヒトの血液に比べて胆管癌患者の血液にモルガネラ属細菌由来小胞が有意に減少していることを確認した（表７および図５参照）。 [Example 6. Metagenomic analysis of vesicles derived from blood bacteria in patients with bile duct cancer]
Morganella was subjected to metagenome analysis by extracting genes from vesicles present in the blood of 79 bile duct cancer patients and 259 normal humans whose age and gender were matched by the method of Example 2. The distribution of vesicles derived from the genus bacteria was evaluated. As a result, it was confirmed that vesicles derived from Morganella spp. Are significantly reduced in the blood of patients with cholangiocarcinoma as compared with the blood of normal humans (see Table 7 and FIG. 5).

Figure 2022028794000008
Figure 2022028794000008

［実施例７．乳癌患者の尿細菌由来小胞メタゲノム分析］
実施例２の方法で乳癌患者１２７人の尿および、性別と年齢をマッチングした正常対照群２２０人の尿を対象に、尿内に存在する小胞から遺伝子を抽出してメタゲノム分析を行った後、モルガネラ属細菌由来小胞の分布を評価した。その結果、正常ヒトの尿に比べて乳癌患者の尿にモルガネラ属細菌由来小胞が有意に減少していることを確認した（表８および図６参照）。 [Example 7. Urine bacteria-derived vesicle metagenomic analysis of breast cancer patients]
After performing metagenome analysis by extracting genes from vesicles present in the urine of 127 breast cancer patients and 220 normal control group urines whose sex and age were matched by the method of Example 2. , The distribution of vesicles derived from Morganella spp. Was evaluated. As a result, it was confirmed that vesicles derived from Morganella spp. Were significantly reduced in the urine of breast cancer patients as compared with the urine of normal humans (see Table 8 and FIG. 6).

Figure 2022028794000009
Figure 2022028794000009

［実施例８．卵巣癌患者の血液および尿細菌由来小胞メタゲノム分析］
実施例２の方法で卵巣癌患者１３７人および、年齢と性別をマッチングした正常ヒト１３９人の血液を対象に、血液内に存在する小胞から遺伝子を抽出してメタゲノム分析を行った後、モルガネラ属細菌由来小胞の分布を評価した。その結果、正常ヒトの血液に比べて卵巣癌患者の血液にモルガネラ属細菌由来小胞が有意に減少していることを確認した（表９および図７ａ参照）。 [Example 8. Metagenomic analysis of vesicles derived from blood and urinary bacteria in patients with ovarian cancer]
Morganella was subjected to metagenome analysis by extracting genes from vesicles present in the blood of 137 ovarian cancer patients and 139 normal humans whose age and gender were matched by the method of Example 2. The distribution of vesicles derived from genus bacteria was evaluated. As a result, it was confirmed that vesicles derived from Morganella spp. Are significantly reduced in the blood of ovarian cancer patients as compared with the blood of normal humans (see Table 9 and FIG. 7a).

Figure 2022028794000010
Figure 2022028794000010

また、実施例２の方法で卵巣癌患者１３６人の尿および、性別と年齢をマッチングした正常対照群１３６人の尿を対象に、尿内に存在する小胞から遺伝子を抽出してメタゲノム分析を行った後、モルガネラ属細菌由来小胞の分布を評価した。その結果、正常ヒトの尿に比べて卵巣癌患者の尿にモルガネラ属細菌由来小胞が有意に減少していることを確認した（表１０および図７ｂ参照）。 In addition, for the urine of 136 ovarian cancer patients and the urine of 136 normal control groups whose sex and age were matched by the method of Example 2, genes were extracted from vesicles present in the urine and metagenome analysis was performed. After that, the distribution of vesicles derived from Morganella spp. Was evaluated. As a result, it was confirmed that vesicles derived from Morganella spp. Were significantly reduced in the urine of ovarian cancer patients as compared with the urine of normal humans (see Table 10 and FIG. 7b).

Figure 2022028794000011
Figure 2022028794000011

［実施例９．膀胱癌患者の血液および尿細菌由来小胞メタゲノム分析］
実施例２の方法で膀胱癌患者９６人の血液および、性別と年齢をマッチングした正常対照群１８４人の血液を対象に、血液内に存在する小胞から遺伝子を抽出してメタゲノム分析を行った後、モルガネラ属細菌由来小胞の分布を評価した。その結果、正常ヒトの血液に比べて膀胱癌患者の血液にモルガネラ属細菌由来小胞が有意に減少していることを確認した（表１１および図８ａ参照）。 [Example 9. Blood and urinary bacterial vesicle metagenomic analysis of bladder cancer patients]
In the blood of 96 patients with bladder cancer and the blood of 184 normal control groups whose sex and age were matched by the method of Example 2, genes were extracted from vesicles present in the blood and metagenome analysis was performed. Later, the distribution of vesicles derived from Morganella spp. Was evaluated. As a result, it was confirmed that vesicles derived from Morganella spp. Are significantly reduced in the blood of bladder cancer patients as compared with the blood of normal humans (see Table 11 and FIG. 8a).

Figure 2022028794000012
Figure 2022028794000012

また、実施例２の方法で膀胱癌患者９５人の尿および、性別と年齢をマッチングした正常対照群１５７人の尿を対象に、尿内に存在する小胞から遺伝子を抽出してメタゲノム分析を行った後、モルガネラ属細菌由来小胞の分布を評価した。その結果、正常ヒトの尿に比べて膀胱癌患者の尿にモルガネラ属細菌由来小胞が有意に減少していることを確認した（表１２および図８ｂ参照）。 In addition, for the urine of 95 bladder cancer patients and the urine of 157 normal control groups whose sex and age were matched by the method of Example 2, genes were extracted from vesicles present in the urine and metagenome analysis was performed. After that, the distribution of vesicles derived from Morganella spp. Was evaluated. As a result, it was confirmed that vesicles derived from Morganella spp. Are significantly reduced in the urine of bladder cancer patients as compared with the urine of normal humans (see Table 12 and FIG. 8b).

Figure 2022028794000013
Figure 2022028794000013

［実施例１０．前立腺癌患者の尿細菌由来小胞メタゲノム分析］
実施例２の方法で前立腺癌患者５３人の尿および、性別と年齢をマッチングした正常対照群１５９人の尿を対象に、尿内に存在する小胞から遺伝子を抽出してメタゲノム分析を行った後、モルガネラ属細菌由来小胞の分布を評価した。その結果、正常ヒトの尿に比べて前立腺癌患者の尿にモルガネラ属細菌由来小胞が有意に減少していることを確認した（表１３および図９参照）。 [Example 10. Urine bacteria-derived vesicle metagenomic analysis of prostate cancer patients]
In the urine of 53 prostate cancer patients and the urine of 159 normal control groups whose sex and age were matched by the method of Example 2, genes were extracted from vesicles present in the urine and metagenome analysis was performed. Later, the distribution of vesicles derived from Morganella spp. Was evaluated. As a result, it was confirmed that vesicles derived from Morganella spp. Are significantly reduced in the urine of prostate cancer patients as compared with the urine of normal humans (see Table 13 and FIG. 9).

Figure 2022028794000014
Figure 2022028794000014

［実施例１１．リンパ腫患者の血液細菌由来小胞メタゲノム分析］
実施例２の方法でリンパ腫患者６３人および、年齢と性別をマッチングした正常ヒト５３人の血液を対象に、血液内に存在する小胞から遺伝子を抽出してメタゲノム分析を行った後、モルガネラ属細菌由来小胞の分布を評価した。その結果、正常ヒトの血液に比べてリンパ腫患者の血液にモルガネラ属細菌由来小胞が有意に減少していることを確認した（表１４および図１０参照）。 [Example 11. Blood bacteria-derived vesicle metagenomic analysis of lymphoma patients]
In the blood of 63 lymphoma patients and 53 normal humans whose age and gender were matched by the method of Example 2, genes were extracted from vesicles present in the blood and metagenome analysis was performed, and then Morganella spp. The distribution of bacterial vesicles was evaluated. As a result, it was confirmed that vesicles derived from Morganella spp. Were significantly reduced in the blood of lymphoma patients as compared with the blood of normal humans (see Table 14 and FIG. 10).

Figure 2022028794000015
Figure 2022028794000015

［実施例１２．心臓疾患患者の血液細菌由来小胞メタゲノム分析］
実施例２の方法で心筋梗塞患者５７人の血液および、性別と年齢をマッチングした正常対照群１６３人の血液を対象に、血液内に存在する小胞から遺伝子を抽出してメタゲノム分析を行った後、モルガネラ属細菌由来小胞の分布を評価した。その結果、正常ヒトの血液に比べて心筋梗塞患者の血液にモルガネラ属細菌由来小胞が有意に減少していることを確認した（表１５および図１１参照）。 [Example 12. Metagenomic analysis of vesicles derived from blood bacteria in patients with heart disease]
In the blood of 57 patients with myocardial infarction and the blood of 163 normal control groups whose sex and age were matched by the method of Example 2, genes were extracted from vesicles present in the blood and metagenome analysis was performed. Later, the distribution of vesicles derived from Morganella spp. Was evaluated. As a result, it was confirmed that vesicles derived from Morganella spp. Were significantly reduced in the blood of patients with myocardial infarction as compared with the blood of normal humans (see Table 15 and FIG. 11).

Figure 2022028794000016
Figure 2022028794000016

また、実施例２の方法で拡張性心筋症患者７２人の血液および、性別と年齢をマッチングした正常対照群１６３人の血液を対象に、血液内に存在する小胞から遺伝子を抽出してメタゲノム分析を行った後、モルガネラ属細菌由来小胞の分布を評価した。その結果、正常ヒトの血液に比べて拡張性心筋症患者の血液にモルガネラ属細菌由来小胞が有意に減少していることを確認した（表１６および図１２参照）。 In addition, the metagenome was obtained by extracting genes from the vesicles present in the blood of 72 patients with dilated cardiomyopathy and the blood of 163 normal control groups whose sex and age were matched by the method of Example 2. After the analysis, the distribution of vesicles derived from Morganella spp. Was evaluated. As a result, it was confirmed that vesicles derived from Morganella spp. Were significantly reduced in the blood of patients with dilated cardiomyopathy as compared with the blood of normal humans (see Table 16 and FIG. 12).

Figure 2022028794000017
Figure 2022028794000017

また、実施例２の方法で心房細動患者３２人の血液と性別と年齢をマッチングした正常対照群３２人の血液を対象に、血液内に存在する小胞から遺伝子を抽出してメタゲノム分析を行った後、モルガネラ属細菌由来小胞の分布を評価した。その結果、正常ヒトの血液に比べて心房細動患者の血液にモルガネラ属細菌由来小胞が有意に減少していることを確認した（表１７および図１３参照）。 In addition, for the blood of 32 patients with atrial fibrillation and the blood of 32 normal control groups whose sex and age were matched by the method of Example 2, genes were extracted from vesicles existing in the blood and metagenome analysis was performed. After that, the distribution of vesicles derived from Morganella spp. Was evaluated. As a result, it was confirmed that vesicles derived from Morganella spp. Are significantly reduced in the blood of patients with atrial fibrillation as compared with the blood of normal humans (see Table 17 and FIG. 13).

Figure 2022028794000018
Figure 2022028794000018

また、実施例２の方法で異型狭心症患者８０人および、年齢と性別をマッチングした正常ヒト８０人の血液を対象に、血液内に存在する小胞から遺伝子を抽出してメタゲノム分析を行った後、モルガネラ属細菌由来小胞の分布を評価した。その結果、正常ヒトの血液に比べて異型狭心症患者の血液にモルガネラ属細菌由来小胞が有意に減少していることを確認した（表１８および図１４参照）。 In addition, for the blood of 80 patients with atypical angina and 80 normal humans whose age and gender were matched by the method of Example 2, genes were extracted from vesicles existing in the blood and metagenome analysis was performed. After that, the distribution of vesicles derived from Morganella spp. Was evaluated. As a result, it was confirmed that vesicles derived from Morganella spp. Were significantly reduced in the blood of patients with atypical angina as compared with the blood of normal humans (see Table 18 and FIG. 14).

Figure 2022028794000019
Figure 2022028794000019

［実施例１３．脳卒中患者の血液細菌由来小胞メタゲノム分析］
実施例２の方法で脳卒中患者１１５人および、年齢と性別をマッチングした正常ヒト１０９人の血液を対象に、血液内に存在する小胞から遺伝子を抽出してメタゲノム分析を行った後、モルガネラ属細菌由来小胞の分布を評価した。その結果、正常ヒトの血液に比べて脳卒中患者の血液にモルガネラ属細菌由来小胞が有意に減少していることを確認した（表１９および図１５参照）。 [Example 13. Blood bacteria-derived vesicle metagenomic analysis of stroke patients]
In the blood of 115 stroke patients and 109 normal humans whose age and gender were matched by the method of Example 2, genes were extracted from vesicles present in the blood and metagenome analysis was performed, and then Morganella spp. The distribution of bacterial vesicles was evaluated. As a result, it was confirmed that vesicles derived from Morganella spp. Were significantly reduced in the blood of stroke patients as compared with the blood of normal humans (see Table 19 and FIG. 15).

Figure 2022028794000020
Figure 2022028794000020

［実施例１４．糖尿病患者の血液細菌由来小胞メタゲノム分析］
実施例２の方法で糖尿病患者７３人の血液および、性別と年齢をマッチングした正常対照群１４６人の血液を対象に、血液内に存在する小胞から遺伝子を抽出してメタゲノム分析を行った後、モルガネラ属細菌由来小胞の分布を評価した。その結果、正常ヒトの血液に比べて糖尿病患者の血液にモルガネラ属細菌由来小胞が有意に減少していることを確認した（表２０および図１６ａ参照）。 [Example 14. Blood bacteria-derived vesicle metagenomic analysis of diabetic patients]
After performing metagenome analysis by extracting genes from vesicles present in the blood of 73 diabetic patients and 146 normal control groups whose sex and age were matched by the method of Example 2. , The distribution of vesicles derived from Morganella spp. Was evaluated. As a result, it was confirmed that vesicles derived from Morganella spp. Were significantly reduced in the blood of diabetic patients as compared with the blood of normal humans (see Table 20 and FIG. 16a).

Figure 2022028794000021
Figure 2022028794000021

また、実施例２の方法で糖尿病患者６０人および、年齢と性別をマッチングした正常ヒト１３４人の尿を対象に、尿内に存在する小胞から遺伝子を抽出してメタゲノム分析を行った後、モルガネラ属細菌由来小胞の分布を評価した。その結果、正常ヒトの尿に比べて糖尿病患者の尿にモルガネラ属細菌由来小胞が有意に減少していることを確認した（表２１および図１６ｂ参照）。 Further, in the urine of 60 diabetic patients and 134 normal humans whose age and sex were matched by the method of Example 2, genes were extracted from vesicles existing in the urine and metagenome analysis was performed. The distribution of vesicles derived from Morganella spp. Was evaluated. As a result, it was confirmed that vesicles derived from Morganella spp. Were significantly reduced in the urine of diabetic patients as compared with the urine of normal humans (see Table 21 and FIG. 16b).

Figure 2022028794000022
Figure 2022028794000022

［実施例１５．パーキンソン病患者の尿細菌由来小胞メタゲノム分析］
実施例２の方法でパーキンソン病患者３９人の尿および、性別と年齢をマッチングした正常対照群７９人の尿を対象に、尿内に存在する小胞から遺伝子を抽出してメタゲノム分析を行った後、モルガネラ属細菌由来小胞の分布を評価した。その結果、正常ヒトの尿に比べてパーキンソン病患者の尿にモルガネラ属細菌由来小胞が有意に減少していることを確認した（表２２および図１７参照）。 [Example 15. Metagenomic analysis of vesicles derived from urinary bacteria in patients with Parkinson's disease]
In the urine of 39 Parkinson's disease patients and the urine of 79 normal control groups whose sex and age were matched by the method of Example 2, genes were extracted from vesicles present in the urine and metagenome analysis was performed. Later, the distribution of vesicles derived from Morganella spp. Was evaluated. As a result, it was confirmed that vesicles derived from Morganella spp. Were significantly reduced in the urine of Parkinson's disease patients as compared with the urine of normal humans (see Table 22 and FIG. 17).

Figure 2022028794000023
Figure 2022028794000023

［実施例１６．モルガネラ・モルガニイ培養液から小胞分離］
前記実施例に基づいて、モルガネラ・モルガニイ（Ｍ．ｍｏｒｇａｎｉｉ）菌株は、韓国微生物保存センター（ＫＣＣＭ）で標準菌株１種（ＭＭＲ１０１）およびヒトから分離した２種の分離菌株（ＭＭＲ２０１、ＭＭＲ２０２）を培養した後、培養液から小胞を分離して特性を分析した。モルガネラ菌株（Ｍ．ｍｏｒｇａｎｉｉ）を３７℃培養器で吸光度（ＯＤ６００）が１．０～１．５になるまでＬＢ（Ｌｕｒｉａ－Ｂｅｒｔａｎｉ）培地で培養した後、サブカルチャーした。以後、菌株が含まれている培養液を回収して１０，０００ｇ、４℃で２０分間遠心分離して菌株を除去し、０．２２μｍのフィルターに濾過した。濾過した上澄み液を１００ｋＤａ Ｐｅｌｌｉｃｏｎ ２ Ｃａｓｓｅｔｔｅフィルターメンブレイン（Ｍｅｒｃｋ Ｍｉｌｌｉｐｏｒｅ，ＵＳ）でＭａｓｔｅｒＦｌｅｘ ｐｕｍｐ ｓｙｓｔｅｍ（Ｃｏｌｅ－Ｐａｒｍｅｒ，ＵＳ）を用いてマイクロフィルター濾過を通じて５０ｍｌ以下の体積に濃縮した。濃縮させた上澄み液をさらに０．２２μｍのフィルターで濾過した。以後、ＢＣＡアッセイを用いてタンパク質を定量し、得られた小胞に対して下記実験を実施した。 [Example 16. Vesicle separation from Morganella-Morganella culture medium]
Based on the above-mentioned Examples, the Morganella morganii strain cultivates one standard strain (MMR101) and two isolated strains (MMR201, MMR202) isolated from humans at the Korea Microbial Conservation Center (KCCM). After that, the vesicles were separated from the culture medium and their characteristics were analyzed. Morganella strain (M. morganii) was cultured in LB (Luria-Bertani) medium until the absorbance (OD600) reached 1.0 to 1.5 in a 37 ° C. incubator, and then subcultured. After that, the culture broth containing the strain was collected and centrifuged at 10,000 g at 4 ° C. for 20 minutes to remove the strain, and filtered through a 0.22 μm filter. The filtered supernatant was concentrated to a volume of 50 ml or less through microfilter filtration using a MasterFlex pump system (Cole-Parmer, US) with a 100 kDa Pellicon 2 Cassette filter membrane (Merck Millipore, US). The concentrated supernatant was further filtered through a 0.22 μm filter. After that, the protein was quantified using the BCA assay, and the following experiment was carried out on the obtained vesicles.

［実施例１７．モルガネラ・モルガニイ由来小胞の細胞死滅効果］
炎症細胞でモルガネラ・モルガニイ由来小胞（Ｍ．ｍｏｒｇａｎｉｉ ＥＶ）の細胞死滅効果を評価するために、マウスマクロファージ細胞株であるＲａｗ ２６４．７細胞にモルガネラ・モルガニイ（ＭＭＲ１０１、ＭＭＲ２０１、ＭＭＲ２０２）由来小胞を多様な濃度（０．１、１、１０μｇ／ｍｌ）で処理した後、細胞死滅程度を評価した。より具体的に、４８ウェル細胞培養プレート内に５×１０４個ずつ分注したＲａｗ ２６４．７細胞にＤＭＥＭ無血清培地で希釈した多様な濃度のモルガネラ・モルガニイ（ＭＭＲ１０１、ＭＭＲ２０１、ＭＭＲ２０２）由来小胞を処理して１２時間の間培養した。以後、細胞死滅は、ＥＺ－ＣＹＴＯＸ（Ｄｏｇｅｎ，Ｋｏｒｅａ）を用いて測定した。その結果、モルガネラ・モルガニイ（ＭＭＲ１０１、ＭＭＲ２０１、ＭＭＲ２０２）由来小胞の処理時に細胞死滅は観察されなかった（図１８参照）。 [Example 17. Cell-killing effect of vesicles derived from Morganella and Morganella]
In order to evaluate the cell-killing effect of Morganella-Morganella-derived vesicles (M. morganii EV) in inflammatory cells, Morganella-Morganella-derived vesicles (MMR101, MMR201, MMR202) -derived vesicles were added to Raw 264.7 cells, which are mouse macrophage cell lines. Was treated at various concentrations (0.1, 1, 10 μg / ml) and then the degree of cell death was evaluated. More specifically, small doses derived from Morganella Morganii (MMR101, MMR201, MMR202) diluted in DMEM serum-free medium in Raw 264.7 cells dispensed in 5 × 10 4 cells in a 48-well cell culture plate. The vesicles were treated and cultured for 12 hours. Subsequently, cell death was measured using EZ-CYTOX (Dogen, Korea). As a result, no cell death was observed during treatment of vesicles derived from Morganella morganii (MMR101, MMR201, MMR202) (see FIG. 18).

［実施例１８．モルガネラ・モルガニイ由来小胞の抗炎症効果］
モルガネラ・モルガニイ由来小胞が、炎症細胞で炎症メディエーターの分泌に対する影響を調べてみるために、マウスマクロファージ細胞株であるＲａｗ ２６４．７細胞にモルガネラ・モルガニイ（ＭＭＲ１０１）由来小胞を多様な濃度（０．１、１、１０μｇ／ｍｌ）で処理した後、炎症疾患病原性小胞である大腸菌由来小胞（Ｅ．ｃｏｌｉ ＥＶ）を処理して炎症メディエーター（ＩＬ－６、ＴＮＦ－αなど）の分泌量を測定した。より具体的に、Ｒａｗ ２６４．７細胞を１ｘ１０５個ずつ２４ウェル細胞培養プレートに分注した後、２４時間の間ＤＭＥＭ完全培地で培養させた。以後、培養上澄み液を１．５ｍｌのチューブに集めて３０００ｇで５分間遠心分離して上澄み液を集めて４℃に保管しておいて、ＥＬＩＳＡ分析を進めた。その結果、モルガネラ・モルガニイ由来小胞を前処理した場合、大腸菌由来小胞によるＩＬ－６およびＴＮＦ－αの分泌が顕著に抑制されることを確認した（図１９ａおよび１９ｂ参照）。特に、モルガネラ・モルガニイ由来小胞を前処理した場合に、マクロファージでＴＮＦ－αの分泌がラクトバチルスプランタルム小胞に比べて抑制程度が顕著に高かった（図１９ｂ参照）。 [Example 18. Anti-inflammatory effect of vesicles derived from Morganella and Morganella]
In order to investigate the effect of Morganella-Morganii-derived vesicles on the secretion of inflammatory mediators in inflammatory cells, various concentrations of Morganella-Morganii (MMR101) -derived vesicles were added to Raw 264.7 cells, which are mouse macrophage cell lines (MMR101). After treatment with 0.1, 1, 10 μg / ml), vesicles derived from E. coli EV, which are pathogenic vesicles of inflammatory diseases, are treated with inflammatory mediators (IL-6, TNF-α, etc.). The amount of secretion was measured. More specifically, Raw 264.7 cells were dispensed into 24-well cell culture plates in groups of 1x10 and 5 cells, and then cultured in DMEM complete medium for 24 hours. After that, the culture supernatant was collected in a 1.5 ml tube, centrifuged at 3000 g for 5 minutes, and the supernatant was collected and stored at 4 ° C. to proceed with the ELISA analysis. As a result, it was confirmed that when the vesicles derived from Morganella and Morganella were pretreated, the secretion of IL-6 and TNF-α by the vesicles derived from Escherichia coli was remarkably suppressed (see FIGS. 19a and 19b). In particular, when vesicles derived from Morganella and Morganella were pretreated, the degree of suppression of TNF-α secretion in macrophages was significantly higher than that in Lactobacillus plantarum vesicles (see FIG. 19b).

また、多様なサンプルから分離したモルガネラ・モルガニイ菌株から分離した小胞の抗炎症効果を評価するために、多様な濃度（０．１、１、１０μｇ／ｍｌ）のモルガネラ・モルガニイ（ＭＭＲ１０１、ＭＭＲ２０１、ＭＭＲ２０２）由来小胞をマウスマクロファージ細胞株に１２時間前処理した後、病原性小胞である大腸菌由来小胞１μｇ／ｍｌを処理し、１２時間後に炎症性サイトカインであるＴＮＦ－αの分泌をＥＬＩＳＡで測定した。その結果、有用微生物対照群であるラクトバチルスプランタルム（Ｌａｃｔｏｂａｃｉｌｌｕｓ ｐｌａｎｔａｒｕｍ）由来小胞の前処理によるＴＮＦ－αの分泌抑制効果よりモルガネラ・モルガニイ（ＭＭＲ１０１、ＭＭＲ２０１、ＭＭＲ２０２）由来小胞の抑制効果がさらに高いことを確認した（図２０参照）。これは、モルガネラ・モルガニイのソースに関係なくモルガネラ・モルガニイ菌が分泌する小胞が、抗炎症効果があることを意味する。 In addition, in order to evaluate the anti-inflammatory effect of vesicles isolated from Morganella Morganii strains isolated from various samples, Morganella Morganii (MMR101, MMR201, MMR101, MMR201, at various concentrations (0.1, 1, 10 μg / ml), MMR202) -derived vesicles are pretreated with mouse macrophage cell lines for 12 hours, then treated with 1 μg / ml of pathogenic vesicles derived from Escherichia coli, and 12 hours later, the secretion of the inflammatory cytokine TNF-α is ELISA. Measured at. As a result, the effect of suppressing the secretion of TNF-α by the pretreatment of the vesicles derived from Lactobacillus plantarum, which is a useful microorganism control group, is further enhanced by the effect of suppressing the vesicles derived from Morganella morganii (MMR101, MMR201, MMR202). It was confirmed that it was high (see FIG. 20). This means that the vesicles secreted by Morganella Morganella, regardless of the source of Morganella Morganella, have anti-inflammatory effects.

［実施例１９．モルガネラ・モルガニイ由来小胞の抗炎症作用に対する熱または酸処理の効果］
前記実施例１８を通じてモルガネラ・モルガニイ標準菌株（ＭＭＲ１０１）および分離菌株（ＭＭＲ２０１）由来小胞の抗炎症効果を確認し、ひいては、前記小胞の安定性および有効物質の特性を具体的に調べてみようとした。このために、１００℃で１０分間煮沸、又は１０分間酸処理（ｐＨ２．０）をした２種のモルガネラ・モルガニイ由来小胞（ＭＭＲ１０１、ＭＭＲ２０１）をマクロファージ（Ｒａｗ ２６４．７）に前処理して抗炎症効果を評価した。その結果、小胞を１００℃の煮沸処理や、酸処理をしても、モルガネラ・モルガニイ由来小胞の抗炎症効果が維持されることを確認した（図２１参照）。これは、モルガネラ・モルガニイ由来小胞の抗炎症作用は、温度と酸に対して安定であることを意味する。 [Example 19. Effect of heat or acid treatment on the anti-inflammatory effect of Morganella-Morganella-derived vesicles]
Through Example 18, the anti-inflammatory effect of vesicles derived from the Morganella-Morganella standard strain (MMR101) and the isolated strain (MMR201) will be confirmed, and the stability of the vesicles and the characteristics of the effective substance will be specifically investigated. And said. For this purpose, two types of Morganella-Morganella-derived vesicles (MMR101, MMR201) boiled at 100 ° C. for 10 minutes or acid-treated for 10 minutes (pH 2.0) were pretreated with macrophages (Raw 264.7). The anti-inflammatory effect was evaluated. As a result, it was confirmed that the anti-inflammatory effect of the vesicles derived from Morganella and Morganella was maintained even when the vesicles were boiled at 100 ° C. or treated with acid (see FIG. 21). This means that the anti-inflammatory effect of Morganella-Morganella-derived vesicles is stable to temperature and acid.

［実施例２０．モルガネラ・モルガニイ由来小胞の抗癌効果］
前記実施例に基づいて、ひいては、モルガネラ・モルガニイ由来小胞の抗癌効果を調べてみようとした。このために、図２２に示されたように、モルガネラ・モルガニイ分離菌株（ＭＭＲ２０１）由来小胞を６週齢Ｃ５７ＢＬ／６雄性マウスに腹腔注射または経口で投与し、投与４日目に癌細胞株（ＣＴ２６ ｃｅｌｌ）を皮下で注射して癌モデルを作成した。癌細胞株を投与した後、モルガネラ・モルガニイ分離菌株由来小胞を腹腔注射または経口で毎日投与し、２４日目まで癌組織の大きさを測定した（図２２参照）。その結果、癌組織の大きさは、対照群である生理食塩水経口投与群に比べて、前記小胞を腹腔注射で投与したマウスと経口で投与したマウスにおいて癌組織の大きさが減少し、特に、経口で投与した場合に大きさがさらに減少した（図２３参照）。これは、モルガネラ・モルガニイ由来小胞を投与したとき、癌組織の成長を効率的に抑制することができることを意味する。 [Example 20. Anti-cancer effect of vesicles derived from Morganella and Morganella]
Based on the above examples, we tried to investigate the anticancer effect of Morganella-Morganella-derived vesicles. To this end, as shown in FIG. 22, vesicles derived from the Morganella Morganii isolate (MMR201) were administered intraperitoneally or orally to 6 week old C57BL / 6 male mice and the cancer cell line was administered on day 4 of administration. (CT26 cell) was injected subcutaneously to create a cancer model. After administration of the cancer cell line, vesicles derived from the Morganella / Morganii isolate were administered intraperitoneally or orally daily, and the size of the cancer tissue was measured until the 24th day (see FIG. 22). As a result, the size of the cancer tissue was reduced in the mice to which the vesicles were intraperitoneally injected and the mice to which the vesicles were orally administered, as compared with the physiological saline oral administration group which was the control group. In particular, the size was further reduced when administered orally (see FIG. 23). This means that when Morganella-Morganella-derived vesicles are administered, the growth of cancerous tissue can be effectively suppressed.

前記記述した本発明の説明は、例示のためのものであり、本発明の属する技術分野における通常の知識を有する者は、本発明の技術的思想や必須の特徴を変更することなく、他の具体的な形態に容易に変形が可能であることを理解することができる。したがって、以上で記述した実施例は、すべての面において例示的なものであり、限定的でないものと理解しなければならない。 The description of the present invention described above is for illustrative purposes only, and a person having ordinary knowledge in the technical field to which the present invention belongs does not change the technical idea or essential features of the present invention. It can be understood that it can be easily transformed into a specific form. Therefore, it should be understood that the examples described above are exemplary in all respects and are not limiting.

本発明によるモルガネラ属細菌由来小胞は、胃癌、大腸癌、すい臓癌、胆管癌、乳癌、卵巣癌、膀胱癌、前立腺癌、リンパ腫、心筋梗塞、心筋症、心房細動、異型狭心症、脳卒中、糖尿病、およびパーキンソン病に対する診断方法、および前記疾患または炎症性疾患に対する食品または薬物などの予防用あるいは治療用組成物に有用に用いられるものと期待される。 The vesicles derived from Morganella spp. According to the present invention include gastric cancer, colon cancer, pancreatic cancer, bile duct cancer, breast cancer, ovarian cancer, bladder cancer, prostate cancer, lymphoma, myocardial infarction, myocardial disease, atrial fibrillation, atypical angina, It is expected to be usefully used in diagnostic methods for stroke, diabetes, and Parkinson's disease, and in prophylactic or therapeutic compositions such as foods or drugs for the diseases or inflammatory diseases.

Claims (13)

モルガネラ（Ｍｏｒｇａｎｅｌｌａ）属細菌由来小胞を有効成分として含む、胃癌、大腸癌、すい臓癌、胆管癌、乳癌、卵巣癌、膀胱癌、前立腺癌、リンパ腫、心筋梗塞、心筋症、心房細動、異型狭心症、脳卒中、糖尿病、パーキンソン病、および炎症性疾患よりなる群から選ばれる１つ以上の疾病の予防または治療用薬学的組成物。 Gastric cancer, colon cancer, pancreatic cancer, bile duct cancer, breast cancer, ovarian cancer, bladder cancer, prostate cancer, lymphoma, myocardial infarction, myocardial disease, atrial fibrillation, atypical, containing vesicles derived from Morganella bacteria as active ingredients A pharmaceutical composition for the prevention or treatment of one or more diseases selected from the group consisting of angina, stroke, diabetes, Parkinson's disease, and inflammatory diseases. 前記炎症性疾患は、アトピー皮膚炎、にきび、乾癬、副鼻腔炎、鼻炎、結膜炎、喘息、皮膚炎、炎症性コラーゲン血管疾患、糸球体腎炎、脳炎、炎症性腸炎、慢性閉鎖性肺疾患、敗血症、敗血症性ショック、肺線維症、未分化脊椎関節症、未分化関節症、関節炎、炎症性骨溶解、ウイルスまたはバクテリア感染による慢性炎症疾患、大腸炎、潰瘍性大腸炎、炎症性腸疾患、関節炎、関節リウマチ、反応性関節炎、骨関節炎、鞏皮症、骨粗しょう症、アテローム性動脈硬化症、心筋炎、心内膜炎、心嚢炎、嚢胞性線維症、橋本甲状腺炎、グレーブス病、ハンセン病、梅毒、ライム病（Ｌｙｍｅ ｄｉｓｅａｓｅ）、ボレリア症（Ｂｏｒｒｅｌｉｏｓｉｓ）、神経性ボレリア症、結核、サルコイドーシス（Ｓａｒｃｏｉｄｏｓｉｓ）、ループス、凍瘡状ループス、結核性ループス、ループス腎炎、全身性エリテマトーデス、黄斑変性、ブドウ膜炎、過敏性腸症候群、クローン病、シェーグレン症候群、線維筋痛、慢性疲労症候群、慢性疲労免疫不全症候群、筋痛性脳脊髄炎、筋萎縮性側索硬化症、パーキンソン病、および多発性硬化症よりなる群から選ばれる１つ以上であることを特徴とする、請求項１に記載の薬学的組成物。 The inflammatory diseases include atopy dermatitis, acne, psoriasis, sinusitis, rhinitis, conjunctivitis, asthma, dermatitis, inflammatory collagen vascular disease, glomerular nephritis, encephalitis, inflammatory enteritis, chronic closed lung disease, and septicemia. , Septic shock, pulmonary fibrosis, undifferentiated lupus erythematosus, undifferentiated arthritis, arthritis, inflammatory osteolysis, chronic inflammatory disease due to viral or bacterial infection, colitis, ulcerative colitis, inflammatory bowel disease, arthritis , Rheumatoid arthritis, reactive arthritis, osteoarthritis, lupus erythematosus, osteoporosis, atherosclerosis, myocarditis, endocarditis, cystitis, cystic fibrosis, Hashimoto thyroiditis, Lyme disease, Hansen's disease, Lupus erythematosus, Lyme disease, Borreliosis, Neuroborreliosis, Tuberculosis, Sarcoidosis, Lupus, Lyme disease, Lupus erythematosus, Lupus erythematosus, Lupus erythematosus, Lupus erythematosus, Lupus erythematosus, Lupus erythematosus, Lupus erythematosus, Lupus erythematosus , Hypersensitivity bowel syndrome, Crohn's disease, Sjogren's syndrome, fibromyalgia, chronic fatigue syndrome, chronic fatigue immunodeficiency syndrome, myopathic encephalomyelitis, my atrophic lateral sclerosis, Parkinson's disease, and multiple sclerosis The pharmaceutical composition according to claim 1, wherein the composition is one or more selected from the group. 前記炎症性疾患は、インターロイキン－６（Ｉｎｔｅｒｌｅｕｋｉｎ－６；ＩＬ－６）または腫瘍壊死因子－アルファ（Ｔｕｍｏｒ ｎｅｃｒｏｓｉｓ ｆａｃｔｏｒ－ａｌｐｈａ；ＴＮＦ－α）により媒介される疾患であることを特徴とする、請求項１又は２に記載の薬学的組成物。 The inflammatory disease is characterized by being a disease mediated by interleukin-6 (IL-6) or tumor necrosis factor-alpha (TNF-α). Item 2. The pharmaceutical composition according to Item 1 or 2. 前記小胞は、平均直径が１０～２００ｎｍであることを特徴とする、請求項１～３のいずれか一項に記載の薬学的組成物。 The pharmaceutical composition according to any one of claims 1 to 3, wherein the vesicle has an average diameter of 10 to 200 nm. 前記小胞は、モルガネラ（Ｍｏｒｇａｎｅｌｌａ）属細菌から自然的または人工的に分泌されることを特徴とする、請求項１～４のいずれか一項に記載の薬学的組成物。 The pharmaceutical composition according to any one of claims 1 to 4, wherein the vesicles are naturally or artificially secreted from a bacterium belonging to the genus Morganella. 前記モルガネラ（Ｍｏｒｇａｎｅｌｌａ）属細菌由来小胞は、モルガネラ・モルガニイ（Ｍｏｒｇａｎｅｌｌａ ｍｏｒｇａｎｉｉ）から分泌されることを特徴とする、請求項１～５のいずれか一項に記載の薬学的組成物。 The pharmaceutical composition according to any one of claims 1 to 5, wherein the vesicles derived from the genus Morganella bacteria are secreted from Morganella morganii. モルガネラ（Ｍｏｒｇａｎｅｌｌａ）属細菌由来小胞を有効成分として含む、胃癌、大腸癌、すい臓癌、胆管癌、乳癌、卵巣癌、膀胱癌、前立腺癌、リンパ腫、心筋梗塞、心筋症、心房細動、異型狭心症、脳卒中、糖尿病、パーキンソン病、および炎症性疾患よりなる群から選ばれる１つ以上の疾病の予防または改善用食品組成物。 Gastric cancer, colon cancer, pancreatic cancer, bile duct cancer, breast cancer, ovarian cancer, bladder cancer, prostate cancer, lymphoma, myocardial infarction, myocardial disease, atrial fibrillation, atypical, containing vesicles derived from Morganella bacteria as active ingredients A food composition for the prevention or amelioration of one or more diseases selected from the group consisting of angina, stroke, diabetes, Parkinson's disease, and inflammatory diseases. 前記炎症性疾患は、アトピー皮膚炎、にきび、乾癬、副鼻腔炎、鼻炎、結膜炎、喘息、皮膚炎、炎症性コラーゲン血管疾患、糸球体腎炎、脳炎、炎症性腸炎、慢性閉鎖性肺疾患、敗血症、敗血症性ショック、肺線維症、未分化脊椎関節症、未分化関節症、関節炎、炎症性骨溶解、ウイルスまたはバクテリア感染による慢性炎症疾患、大腸炎、潰瘍性大腸炎、炎症性腸疾患、関節炎、関節リウマチ、反応性関節炎、骨関節炎、鞏皮症、骨粗しょう症、アテローム性動脈硬化症、心筋炎、心内膜炎、心嚢炎、嚢胞性線維症、橋本甲状腺炎、グレーブス病、ハンセン病、梅毒、ライム病（Ｌｙｍｅ ｄｉｓｅａｓｅ）、ボレリア症（Ｂｏｒｒｅｌｉｏｓｉｓ）、神経性ボレリア症、結核、サルコイドーシス（Ｓａｒｃｏｉｄｏｓｉｓ）、ループス、凍瘡状ループス、結核性ループス、ループス腎炎、全身性エリテマトーデス、黄斑変性、ブドウ膜炎、過敏性腸症候群、クローン病、シェーグレン症候群、線維筋痛、慢性疲労症候群、慢性疲労免疫不全症候群、筋痛性脳脊髄炎、筋萎縮性側索硬化症、パーキンソン病、および多発性硬化症よりなる群から選ばれる１つ以上であることを特徴とする、請求項７に記載の食品組成物。 The inflammatory diseases include atopy dermatitis, acne, psoriasis, sinusitis, rhinitis, conjunctivitis, asthma, dermatitis, inflammatory collagen vascular disease, glomerular nephritis, encephalitis, inflammatory enteritis, chronic closed lung disease, and septicemia. , Septic shock, pulmonary fibrosis, undifferentiated lupus erythematosus, undifferentiated arthritis, arthritis, inflammatory osteolysis, chronic inflammatory disease due to viral or bacterial infection, colitis, ulcerative colitis, inflammatory bowel disease, arthritis , Rheumatoid arthritis, reactive arthritis, osteoarthritis, lupus erythematosus, osteoporosis, atherosclerosis, myocarditis, endocarditis, cystitis, cystic fibrosis, Hashimoto thyroiditis, Lyme disease, Hansen's disease, Lupus erythematosus, Lyme disease, Borreliosis, Neuroborreliosis, Tuberculosis, Sarcoidosis, Lupus, Lyme disease, Lupus erythematosus, Lupus erythematosus, Lupus erythematosus, Lupus erythematosus, Lupus erythematosus, Lupus erythematosus, Lupus erythematosus, Lupus erythematosus , Hypersensitivity bowel syndrome, Crohn's disease, Sjogren's syndrome, fibromyalgia, chronic fatigue syndrome, chronic fatigue immunodeficiency syndrome, myopathic encephalomyelitis, my atrophic lateral sclerosis, Parkinson's disease, and multiple sclerosis The food composition according to claim 7, wherein the food composition is one or more selected from the group. 前記炎症性疾患は、インターロイキン－６（Ｉｎｔｅｒｌｅｕｋｉｎ－６；ＩＬ－６または腫瘍壊死因子－アルファ（Ｔｕｍｏｒ ｎｅｃｒｏｓｉｓ ｆａｃｔｏｒ－ａｌｐｈａ；ＴＮＦ－α）により媒介される疾患であることを特徴とする、請求項７又は８に記載の食品組成物。 The inflammatory disease is characterized by being a disease mediated by interleukin-6 (IL-6 or Tumor necrosis factor-alpha; TNF-α). The food composition according to 7 or 8. 前記小胞は、平均直径が１０～２００ｎｍであることを特徴とする、請求項７～９のいずれか一項に記載の食品組成物。 The food composition according to any one of claims 7 to 9, wherein the vesicles have an average diameter of 10 to 200 nm. 前記小胞は、モルガネラ（Ｍｏｒｇａｎｅｌｌａ）属細菌から自然的または人工的に分泌されることを特徴とする、請求項７～１０のいずれか一項に記載の食品組成物。 The food composition according to any one of claims 7 to 10, wherein the vesicles are naturally or artificially secreted from a bacterium belonging to the genus Morganella. 前記モルガネラ（Ｍｏｒｇａｎｅｌｌａ）属細菌由来小胞は、モルガネラ・モルガニイ（Ｍｏｒｇａｎｅｌｌａ ｍｏｒｇａｎｉｉ）から分泌されることを特徴とする、請求項７～１１のいずれか一項に記載の食品組成物。 The food composition according to any one of claims 7 to 11, wherein the vesicles derived from the genus Morganella bacteria are secreted from Morganella morganii. モルガネラ（Ｍｏｒｇａｎｅｌｌａ）属細菌由来小胞の、胃癌、大腸癌、すい臓癌、胆管癌、乳癌、卵巣癌、膀胱癌、前立腺癌、リンパ腫、心筋梗塞、心筋症、心房細動、異型狭心症、脳卒中、糖尿病、パーキンソン病、および炎症性疾患よりなる群から選ばれる１つ以上の疾病の予防または治療用薬剤の製造のための使用。
Gastric cancer, colon cancer, pancreatic cancer, bile duct cancer, breast cancer, ovarian cancer, bladder cancer, prostate cancer, lymphoma, myocardial infarction, myocardial disease, atrial fibrillation, atypical angina, Use for the manufacture of prophylactic or therapeutic agents for one or more diseases selected from the group consisting of stroke, diabetes, Parkinson's disease, and inflammatory diseases.
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